Funding, Deals & Partnerships: BIOLOGICS & MEDICAL DEVICES; BioMed e-Series; Medicine and Life Sciences Scientific Journal – http://PharmaceuticalIntelligence.com
This discussion that completes and is an epicrisis (summary and critical evaluation) of the series of discussions that preceded it.
Innervation of Heart and Heart Rate
Action of hormones on the circulation
Allogeneic Transfusion Reactions
Graft-versus Host reaction
Unique problems of perinatal period
High altitude sickness
Deep water adaptation
Heart-Lung-and Kidney
Acute Lung Injury
The concept inherent in this series is that the genetic code is an imprint that is translated into a message. It is much the same as a blueprint, or a darkroom photographic image that has to be converted to a print. It is biologically an innovation of evolutionary nature because it establishes a simple and reproducible standard for the transcription of the message through the transcription of the message using strings of nucleotides (oligonucleotides) that systematically transfer the message through ribonucleotides that communicate in the cytoplasm with the cytoskeleton based endoplasmic reticulum (ER), composing a primary amino acid sequence. This process is a quite simple and convenient method of biological activity. However, the simplicity ends at this step. The metabolic components of the cell are organelles consisting of lipoprotein membranes and a cytosol which have particularly aligned active proteins, as in the inner membrane of the mitochondrion, or as in the liposome or phagosome, or the structure of the ER, each of which is critical for energy transduction and respiration, in particular, for the mitochondria, cellular remodeling or cell death, with respect to the phagosome, and construction of proteins with respect to the ER, and anaerobic glycolysis and the hexose monophosphate shunt in the cytoplasmic domain. All of this refers to structure and function, not to leave out the membrane assigned transport of inorganic, and organic ions (electrolytes and metabolites).
I have identified a specific role of the ER, the organelles, and cellular transactions within and between cells that is orchestrated. But what I have outlined is a somewhat limited and rigid model that does not reach into the dynamics of cellular transactions. The DNA has expression that may be old, no longer used messages, and this is perhaps only part of a significant portion of “dark matter”. There is also nuclear DNA that is enmeshed with protein, mRNA that is a copy of DNA, and mDNA is copied to ribosomal RNA (rRNA). There is also rDNA. The classic model is DNA to RNA to protein. However, there is also noncoding RNA, which plays an important role in regulation of transcription.
This has been discussed in other articles. But the important point is that proteins have secondary structure through disulfide bonds, which is determined by position of sulfur amino acids, and by van der Waal forces, attraction and repulsion. They have tertiary structure, which is critical for 3-D structure. When like subunits associate, or dissimilar oligomers, then you have heterodimers and oligomers. These constructs that have emerged over time interact with metabolites within the cell, and also have an important interaction with the extracellular environment.
When you take this into consideration then a more complete picture emerges. The primitive cell or the multicellular organism lives in an environment that has the following characteristics – air composition, water and salinity, natural habitat, temperature, exposure to radiation, availability of nutrients, and exposure to chemical toxins or to predators. In addition, there is a time dimension that proceeds from embryonic stage to birth in mammals, a rapid growth phase, a tapering, and a decline. The time span is determined by body size, fluidity of adaptation, and environmental factors. This is covered in great detail in this work. The last two pieces are in the writing stage that completes the series. Much content has already be presented in previous articles.
The function of the heart, kidneys and metabolism of stressful conditions have already been extensively covered in http://pharmaceuticalintelligence.com in the following and more:
The Amazing Structure and Adaptive Functioning of the Kidneys: Nitric Oxide – Part I
Biochemical Insights of Dr. Jose Eduardo de Salles Roselino
How is it that developments late in the 20th century diverted the attention of
biological processes from a dynamic construct involving interacting chemical
reactions under rapidly changing external conditions effecting tissues and cell
function to a rigid construct that is determined unilaterally by the genome
construct, diverting attention from mechanisms essential for seeing the complete
cellular construct?
Larry, I assume that in case you read the article titled Neo – Darwinism, The
Modern Synthesis and Selfish Genes that bares no relationship with Physiology
with Molecular Biology J. Physiol 2011; 589(5): 1007-11 by Denis Noble, you might
find that it was the key factor required in order to understand the dislodgment
of physiology as a foundation of medical reasoning. In the near unilateral emphasis
of genomic activity as a determinant of cellular activity all of the required general
support for the understanding of my reasoning. The DNA to protein link goes
from triplet sequence to amino acid sequence. That is the realm of genetics.
Further, protein conformation, activity and function requires that environmental
and micro-environmental factors should be considered (Biochemistry). If that
were not the case, we have no way to bridge the gap between the genetic
code and the evolution of cells, tissues, organs, and organisms.
Consider this example of hormonal function. I would like to stress in
the cAMP dependent hormonal response, the transfer of information
that occurs through conformation changes after protein interactions.
This mechanism therefore, requires that proteins must not have their
conformation determined by sequence alone.
Regulatory protein conformation is determined by its sequence plus
the interaction it has in its micro-environment. For instance, if your
scheme takes into account what happens inside the membrane and
that occurs before cAMP, then production is increased by hormone
action. A dynamic scheme will show an effect initially, over hormone
receptor (hormone binding causing change in its conformation) followed
by GTPase change in conformation caused by receptor interaction and
finally, Adenylate cyclase change in conformation and in activity after
GTPase protein binding in a complex system that is dependent on self-
assembly and also, on changes in their conformation in response to
hormonal signals (see R. A Kahn and A. G Gilman 1984 J. Biol. Chem.
v. 259,n 10 pp6235-6240. In this case, trimeric or dimeric G does not
matter). Furthermore, after the step of cAMP increased production we
also can see changes in protein conformation. The effect of increased
cAMP levels over (inhibitor protein and protein kinase protein complex)
also is an effect upon protein conformation. Increased cAMP levels led
to the separation of inhibitor protein (R ) from cAMP dependent protein
kinase (C ) causing removal of the inhibitor R and the increase in C activity.
R stands for regulatory subunit and C for catalytic subunit of the protein
complex.
This cAMP effect over the quaternary structure of the enzyme complex
(C protein kinase + R the inhibitor) may be better understood as an
environmental information producing an effect in opposition to
what may be considered as a tendency towards a conformation
“determined” by the genetic code. This “ideal” conformation
“determined” by the genome would be only seen in crystalline
protein. In carbohydrate metabolism in the liver the hormonal signal
causes a biochemical regulatory response that preserves homeostatic
levels of glucose (one function) and in the muscle, it is a biochemical
regulatory response that preserves intracellular levels of ATP (another
function).
Therefore, sequence alone does not explain conformation, activity
and function of regulatory proteins. If this important regulatory
mechanism was not ignored, the work of S. Prusiner (Prion diseases
and the BSE crisis Stanley B. Prusiner 1997 Science; 278: 245 – 251,
10 October) would be easily understood. We would be accustomed
to reason about changes in protein conformation caused by protein
interaction with other proteins, lipids, small molecules and even ions.
In case this wrong biochemical reasoning is used in microorganisms.
Still it is wrong but, it will cause a minor error most of the time, since
we may reduce almost all activity of microorganism´s proteins to a
single function – The production of another microorganism. However,
even microorganisms respond differently to their micro-environment
despite a single genome (See M. Rouxii dimorphic fungus works,
later). The reason for the reasoning error is, proteins are proteins
and DNA are DNA quite different in chemical terms. Proteins must
change their conformation to allow for fast regulatory responses and
DNA must preserve its sequence to allow for genetic inheritance.
The following article is a review of the central relationship between the action of
metformin as a diabetic medication and its relationship to AMPK, the important and
essential regulator of glucose and lipid metabolism under normal activity, stress, with
its effects on skeletal muscle, the liver, the action of T3 and more.
We start with a case study and a publication in the J Can Med Assoc. Then we shall look
into key literature on these metabolic relationships.
Part I. Metformin , Diabetes Mellitus, and Thyroid Function
Hypothyroidism, Insulin resistance and Metformin
May 30, 2012 By Janie Bowthorpe
The following was written by a UK hypothyroid patient’s mother –
Sarah Wilson.
My daughter’s epilepsy is triggered by unstable blood sugars. And since taking
Metformin to control her blood sugar, she has significantly reduced the number of seizures. I have been doing research and read numerous academic medical journals,
which got me thinking about natural thyroid hormone and Hypothyroidism. My hunch
was that when patients develop hypothyroid symptoms, they are actually becoming
insulin resistant (IR). There are many symptoms in common between women with
polycystic ovaries and hypothyroidism–the hair loss, the weight gain, etc.
(http://insulinhub.hubpages.com/hub/PCOS-and-Hypothyroidism).
A hypothyroid person’s body behaves as if it’s going into starvation mode and so, to
preserve resources and prolong life, the metabolism changes. If hypothyroid is prolonged
or pronounced, then perhaps, chemical preservation mode becomes permanent even
with the reintroduction of thyroid hormones. To get back to normal, they need
a “jump-start” reinitiate a higher rate of metabolism. The kick start is initiated through
AMPK, which is known as the “master metabolic regulating enzyme.”
(http://en.wikipedia.org/wiki/AMP-activated protein kinase).
Note the following comments/partial statements:
“Hypothyroidism is characterized by decreased insulin responsiveness”;
“the pivotal regulatory role of T3 in major metabolic pathways”.
The community knows that T3/NTH (natural thyroid hormone [Armour]) makes
hypothyroid patients feel better – but the medical establishment is averse to T3/NTH
(treating subclinical hypoT (T3/T4 euthyroid) with natural dessicated thyroid (NDT).
The medical establishment might find an alternative view about impaired metabolism
more if shown real proof that the old NDT **was/is** having the right result –i.e., the
T3 is jump-starting the metabolism by re-activatingAMPK.
If NDT also can be used for hypothyroidism without the surmised “dangers” of NTH,
then they should consider it. [The reality in the choice is actually recombinant TH
(Synthroid)]. Metformin is cheap, stable and has very few serious side effects. I use the
car engine metaphor, and refer to glucose as our petrol, AMPK as the spark plug and
both T3 and Metformin as the ignition switches. Sometimes if you have flat batteries in
the car, it doesn’t matter how much you turn the ignition switch or pump the petrol
pedal, all it does is flatten the battery and flood the engine.
Dr. Skinner in the UK has been treating “pre-hypothyroidism” the way that some
doctors treat “pre-diabetes”. Those hypothyroid patients who get treated early
might not have had their AMPK pathways altered and the T4-T3 conversion still works.
There seems to be no reason why thyroid hormone replacement therapy shouldn’t
logically be given to ward off a greater problem down the line.
It’s my belief that there is clear and abundant academic evidence that the AMPK/
Metformin research should branch out to also look at thyroid disease.
Metformin Linked to Risk of Low Levels of Thyroid Hormone
CMAJ (Canadian Medical Association Journal) 09/22/2014
Metformin, the drug commonly for treating type 2 diabetes,
is linked to an increased risk of low thyroid-stimulating hormone (TSH) levels
in patients with underactive thyroids (hypothyroidism),
according to a study in CMAJ (Canadian Medical Association Journal).
Metformin is used to lower blood glucose levels
by reducing glucose production in the liver.
previous studies have raised concerns that
metformin may lower thyroid-stimulating hormone levels.
Study characteristics:
Retrospective long-term
74 300 patient who received metformin and sulfonylurea
25-year study period.
5689 had treated hypothyroidism
59 937 had normal thyroid function.
Metformin and low levels of thyroid-stimulating hormone in patients with type 2 diabetes mellitus
Jean-Pascal Fournier, Hui Yin, Oriana Hoi Yun Yu, Laurent Azoulay +
Centre for Clinical Epidemiology (Fournier, Yin, Yu, Azoulay), Lady Davis Institute,
Jewish General Hospital; Department of Epidemiology, Biostatistics and Occupational
Health (Fournier), McGill University; Division of Endocrinology (Yu), Jewish General
Hospital; Department of Oncology (Azoulay), McGill University, Montréal, Que., Cananda
metformin may lower thyroid-stimulating hormone (TSH) levels.
Objective:
determine whether the use of metformin monotherapy, when compared with
sulfonylurea monotherapy,
is associated with an increased risk of low TSH levels(< 0.4 mIU/L)
in patients with type 2 diabetes mellitus.
Methods:
Used the Clinical Practice Research Datalink,
identified patients who began receiving metformin or sulfonylurea monotherapy
between Jan. 1, 1988, and Dec. 31, 2012.
2 subcohorts of patients with treated hypothyroidism or euthyroidism,
followed them until Mar. 31, 2013.
Used Cox proportional hazards models to evaluate the association of low TSH
levels with metformin monotherapy, compared with sulfonylurea monotherapy,
in each subcohort.
Results:
5689 patients with treated hypothyroidism and 59 937 euthyroid patients were
included in the subcohorts.
For patients with treated hypothyroidism:
495 events of low TSH levels were observed (incidence rate 0.1197/person-years).
322 events of low TSH levels were observed (incidence rate 0.0045/person-years)
in the euthyroid group.
metformin monotherapy was associated with a 55% increased risk of low TSH
levels in patients with treated hypothyroidism (incidence rate 0.0795/person-years
vs.0.1252/ person-years, adjusted hazard ratio [HR] 1.55, 95% confidence
interval [CI] 1.09– 1.20), compared with sulfonylurea monotherapy,
the highest risk in the 90–180 days after initiation (adjusted HR 2.30, 95% CI
1.00–5.29).
No association was observed in euthyroid patients (adjusted HR 0.97, 95% CI 0.69–1.36).
Interpretation: The clinical consequences of this needs further investigation.
Crude and adjusted hazard ratios for suppressed thyroid-stimulating hormone
levels (< 0.1 mIU/L) associated with the use metformin monotherapy, compared
with sulfonylurea monotherapy, in patients with treated hypothyroidism or
euthyroidism and type 2 diabetes
Variable
No. events
suppressed
TSH levels
Person-years of
exposure
Incidence rate,
per 1000 person-years (95% CI)
Crude
HR
Adjusted HR*(95% CI)
Patients with treated hypothyroidism, n = 5689
Sulfonylure, n = 762
18
503
35.8
(21.2–56.6)
1.00
1.00
(reference)
Metformin, n = 4927
130
3 633
35.8
(29.9–42.5)
1.05
0.99
(0.57–1.72)
Euthyroid patients, n = 59 937
Sulfonylurea, n = 7980
12
8 576
1.4
(0.7–2.4)
1.00
1.00
(reference)
Metformin, n = 51 957
75
63 047
1.2
(0.9–1.5)
0.85
1.03
(0.52–2.03)
Part II. Metabolic Underpinning
(Source: Wikipedia, AMPK and thyroid)
5′ AMP-activated protein kinase or AMPK or 5′ adenosine monophosphate-activated protein kinase
is an enzyme that plays a role in cellular energy homeostasis.
It consists of three proteins (subunits) that
together make a functional enzyme, conserved from yeast to humans.
It is expressed in a number of tissues, including the liver, brain, and skeletal
muscle.
The net effect of AMPK activation is stimulation of
hepatic fatty acid oxidation and ketogenesis,
inhibition of cholesterol synthesis,
lipogenesis, and triglyceride synthesis,
inhibition of adipocyte lipolysis and lipogenesis,
stimulation of skeletal muscle fatty acid oxidation and muscle
glucose uptake, and
modulation of insulin secretion by pancreatic beta-cells.
The heterotrimeric protein AMPK is formed by α, β, and γ subunits. Each of these three
subunits takes on a specific role in both the stability and activity of AMPK.
the γ subunit includes four particular Cystathionine beta synthase (CBS) domains
giving AMPK its ability to sensitively detect shifts in the AMP:ATP ratio.
The four CBS domains create two binding sites for AMP commonly referred to as
Bateman domains. Binding of one AMP to a Bateman domain cooperatively
increases the binding affinity of the second AMP to the other Bateman domain.
As AMP binds both Bateman domains the γ subunit undergoes a conformational
change which exposes the catalytic domain found on the α subunit.
It is in this catalytic domain where AMPK becomes activated when
phosphorylation takes place at threonine-172by an upstream AMPK kinase
(AMPKK). The α, β, and γ subunits can also be found in different isoforms.
AMPK acts as a metabolic master switch regulating several intracellular systems
the cellular uptake of glucose,
the β-oxidation of fatty acids and
the biogenesis of glucose transporter 4 (GLUT4) and
mitochondria
The energy-sensing capability of AMPK can be attributed to
its ability to detect and react to fluctuations in the AMP:ATP ratio that take
place during rest and exercise (muscle stimulation).
During muscle stimulation,
AMP increases while ATP decreases, which changes AMPK into a good substrate
for activation.
AMPK activity increases while the muscle cell experiences metabolic stress
brought about by an extreme cellular demand for ATP.
Upon activation, AMPK increases cellular energy levels by
inhibiting anabolic energy consuming pathways (fatty acid synthesis,
protein synthesis, etc.) and
A recent JBC paper on mice at Johns Hopkins has shown that when the activity of brain
AMPK was pharmacologically inhibited,
the mice ate less and lost weight.
When AMPK activity was pharmacologically raised (AICAR see below)
the mice ate more and gained weight.
Research in Britain has shown that the appetite-stimulating hormone ghrelin also
affects AMPK levels.
The antidiabetic drug metformin (Glucophage) acts by stimulating AMPK, leading to
reduced glucose production in the liver and
reduced insulin resistance in the muscle.
(Metformin usually causes weight loss and reduced appetite, not weight gain and
increased appetite, ..opposite of expected from the Johns Hopkins mouse study results.)
Triggering the activation of AMPK can be carried out provided two conditions are met.
an AMP analogthat has been shown to activate AMPK.
Recent LKB1 knockout studies have shown that without LKB1,
electrical and AICAR stimulation of muscleresults in very little
phosphorylation of AMPK and of ACC, providing evidence that
LKB1-STRAD-MO25 is the major AMPKK in muscle.
Two particular adipokines, adiponectin and leptin, have even been demonstrated
to regulate AMPK. A main functions of leptin in skeletal muscle is
the upregulation of fatty acid oxidation.
Leptin works by way of the AMPK signaling pathway, and adiponectin also
stimulates the oxidation of fatty acids via the AMPK pathway, and
Adiponectin also stimulates the uptake of glucose in skeletal muscle.
An increase in enzymes which specialize in glucose uptake in cells such as GLUT4
and hexokinase II are thought to be mediated in part by AMPK when it is activated.
Increases in AMPK activity are brought about by increases in the AMP:ATP ratio
during single bouts of exercise and long-term training.
One of the key pathways in AMPK’s regulation of fatty acid oxidation is the
phosphorylation and inactivation of acetyl-CoA carboxylase.
Acetyl-CoA carboxylase (ACC) converts acetyl-CoA (ACA) to malonyl-CoA
(MCA), an inhibitor of carnitine palmitoyltransferase 1 (CPT-1).
CPT-1 transports fatty acids into the mitochondria for oxidation.
Inactivation of ACC results in increased fatty acid transport and oxidation.
the AMPK induced ACC inactivation and reduced conversion to MCA
may occur as a result of malonyl-CoA decarboxylase (MCD)
MCD as an antagonist to ACC, decarboxylatesmalonyl-CoA to acetyl-CoA
(reversal of ACC conversion of ACA to MCA)
This resultsin decreased malonyl-CoA and increased CPT-1 and fatty acid oxidation.
AMPK also plays an important role in lipid metabolism in the liver. It has long been
known that hepatic ACC has been regulated in the liver.
It phosphorylates and inactivates 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR)
acetyl-CoA(ACA) is converted to mevalonic acid (MVA) by ACC
with inhibition of CPT-1
HMGR converts 3-hydroxy-3-methylglutaryl-CoA, which is made from MVA
which then travels down several more metabolic steps to become cholesterol.
Insulin facilitates the uptake of glucose into cells via increased expression and
translocation of glucose transporter GLUT-4. In addition, glucose is phosphorylated
by hexokinase wheni iot enters the cell. The phosphorylated form keeps glucose from
leaving the cell,
The decreasedthe concentration of glucose molecules creates a gradient for more
glucose to be transported into the cell.
AMPK and thyroid hormone regulate some similar processes. Knowing these similarities,
Winder and Hardie et al. designed an experiment to see if AMPK was influenced by thyroid
hormone. They found that all of the subunits of AMPK were increased in skeletal muscle,
especially in the soleus and red quadriceps, with thyroid hormone treatment. There was
also an increase in phospho-ACC, a marker of AMPK activity.
Winder WW, Hardie DG (July 1999). “AMP-activated protein kinase,
a metabolic master switch: possible roles in type 2 diabetes”. J. Physiol.277
(1 Pt 1): E1–10. PMID10409121.
Winder WW, Hardie DG (February 1996). “Inactivation of acetyl-CoA
carboxylase and activation of AMP-activated protein kinase in muscle
during exercise”. J. Physiol.270 (2 Pt 1): E299–304. PMID8779952.
Hutber CA, Hardie DG, Winder WW (February 1997). “Electrical stimulation
inactivates muscle acetyl-CoA carboxylase and increases AMP-activated
protein kinase”. Am. J. Physiol. 272 (2 Pt 1): E262–6. PMID 9124333
Durante PE, Mustard KJ, Park SH, Winder WW, Hardie DG (July 2002).
“Effects of endurance training on activity and expression of AMP-activated
protein kinase isoforms in rat muscles”. Am. J. Physiol. Endocrinol.
Metab. 283 (1): E178–86. doi:10.1152/ajpendo.00404.2001. PMID 12067859
Corton JM, Gillespie JG, Hardie DG (April 1994). “Role of the AMP-activated
protein kinase in the cellular stress response”. Curr. Biol. 4 (4):
315–24. doi:10.1016/S0960-9822(00)00070-1. PMID 7922340
Winder WW (September 2001). “Energy-sensing and signaling by
AMP-activated protein kinase in skeletal muscle”. J. Appl. Physiol. 91 (3):
1017–28. PMID 11509493
Suter M, Riek U, Tuerk R, Schlattner U, Wallimann T, Neumann D (October
2006). “Dissecting the role of 5′-AMP for allosteric stimulation, activation,
and deactivation of AMP-activated protein kinase”. J. Biol. Chem.
281 (43): 32207–6. doi:10.1074/jbc.M606357200. PMID 16943194
Part III. Pituitary-thyroid axis and diabetes mellitus The Interface Between Thyroid and Diabetes Mellitus
Leonidas H. Duntas, Jacques Orgiazzi, Georg Brabant Clin Endocrinol. 2011;75(1):1-9.
Interaction of Metformin and Thyroid Function
Metformin acts primarily by
suppressing hepatic gluconeogenesis via activation of AMPK
It has the opposite effects on hypothalamic AMPK,
inhibiting activity of the enzyme.
the metformin effects on hypothalamic AMPK activity will
counteractT3 effects at the hypothalamic level.
AMPK therefore represents a direct target for dual regulation
in the hypothalamic partitioning of energy homeostasis.
metformin crossesthe blood–brain barrier and
levels in the pituitary gland are substantially increased.
It convincinglysuppressesTSH
A recent study recruiting 66 patients with benign thyroid nodules furthermore
demonstrated that metformin significantly decreases nodule size in patients with
insulin resistance.[76] The effect of metformin, which was produced over a
6-month treatment period, parallelled a fall in TSH concentrations and achieved a
shrinkage amounting to 30% of the initial nodule size when metformin was
administered alone and up to 55% when it was added to ongoing LT4 treatment.
These studies reveal a
suppressive effect of metformin on TSH secretion patterns in
hypothyroid patients, an effect that is apparently
independent of T4 treatment and does not alter the TH profile.
A rebound of TSH secretion occurs at about 3 months following metformin
withdrawal.
It appears that recommendations for more frequent testing, on an annual to
biannual basis, seems justified in higher risk groups like patients over 50 or 55,
particularly with suggestive symptoms, raised antibody titres or dylipidaemia.
We thus would support the suggestion of an initial TSH and TPO antibody testing
which, as discussed, will help to predict the development of hypothyroidism in
patients with diabetes.
Hypothalamic AMPK and fatty acid metabolism mediate thyroid
regulation of energy balance
M López, L Varela, MJ Vázquez, S Rodríguez-Cuenca, CR González, …, & Vidal-Puig
Nature Medicine 29 Aug 2010; 16: 1001–1008 http://dx.doi.org:/10.1038/nm.2207
Thyroid hormones have widespread cellular effects; however it is unclear whether
their effects on the central nervous system (CNS) contribute to global energy balance.
Here we demonstrate that either
whole-body hyperthyroidism or central administration of triiodothyronine
(T3) decreases
the activity of hypothalamic AMP-activated protein kinase (AMPK),
increases sympathetic nervous system (SNS) activity and
upregulates thermogenic markers in brown adipose tissue (BAT).
Inhibition of the lipogenic pathway in the ventromedial nucleus of the hypothalamus
(VMH) prevents CNS-mediated activation of BAT by thyroid hormone and reverses
the weight loss associated with hyperthyroidism. Similarly, inhibition of thyroid
hormone receptors in the VMH reverses the weight loss associated with hyperthyroidism.
This regulatory mechanism depends on AMPK inactivation, as genetic inhibition of this
enzyme in the VMH of euthyroid rats induces feeding-independent weight loss and
increases expression of thermogenic markers in BAT. These effects are reversed by
pharmacological blockade of the SNS. Thus, thyroid hormone–induced modulation
of AMPK activity and lipid metabolism in the hypothalamus is a major regulator of
whole-body energy homeostasis.
Metabolic Basis for Thyroid Hormone Liver Preconditioning:
Upregulation of AMP-Activated Protein Kinase Signaling
LA Videla,1 V Fernández, P Cornejo, and R Vargas
1Molecular and Clinical Pharmacology Program, Institute of Biomedical Sciences,
Faculty of Medicine, University of Chile, 2Faculty of Medicine, Diego Portales University,
Santiago, Chile
Academic Editors: H. M. Abu-Soud and D. Benke The Scientific World Journal 2012; 2012, ID 475675, 10 pp http://dx.doi.org/10.1100/2012/475675
The liver is a major organ responsible for most functions of cellular metabolism and
a mediator between dietary and endogenous sources of energy for extrahepatic tissues.
In this context, adenosine-monophosphate- (AMP-) activated protein kinase (AMPK)
constitutes an intrahepatic energy sensor
regulating physiological energy dynamics by limiting anabolism and stimulating
catabolism, thus increasing ATP availability.
This is achieved by mechanisms involving direct allosteric activation and
reversible phosphorylation of AMPK, in response to signals such as
energy status,
serum insulin/glucagon ratio,
nutritional stresses,
pharmacological and natural compounds, and
oxidative stress status.
Reactive oxygen species (ROS) lead to cellular AMPK activation and
downstream signaling under several experimental conditions.
Thyroid hormone (L-3,3′,5-triiodothyronine, T3) administration, a condition
that enhances liver ROS generation,
triggers the redox upregulation of cytoprotective proteins
affording preconditioning against ischemia-reperfusion (IR) liver injury.
Data discussed in this work suggest that T3-induced liver activation of AMPK
may be of importance in the promotion of metabolic processes
favouring energy supply for the induction and operation of preconditioning
mechanisms.
These include
antioxidant,
antiapoptotic, and
anti-inflammatory mechanisms,
repair or resynthesis of altered biomolecules,
induction of the homeostatic acute-phase response, and
stimulation of liver cell proliferation,
which are required to cope with the damaging processes set in by IR.
The liver functions as a mediator between dietary and endogenous sources
of energy and extrahepatic organs that continuously require energy, mainly
the brain and erythrocytes, under cycling conditions between fed and fasted states.
In the fed state, where insulin action predominates, digestion-derived glucose is
converted to pyruvate via glycolysis, which is oxidized to produce energy, whereas
fatty acid oxidation is suppressed. Excess glucose can be either stored as hepatic
glycogen or channelled into de novo lipogenesis.
In the fasted state, considerable liver fuel metabolism changes occur due to decreased
serum insulin/glucagon ratio, with higher glucose production as a consequence of
stimulated glycogenolysis and gluconeogenesis (from alanine, lactate, and glycerol).
Major enhancement in fatty acid oxidation also occurs to provide energy for liver
processes and ketogenesis to supply metabolic fuels for extrahepatic tissues. For these
reasons, the liver is considered as the metabolic processing organ of the body, and
alterations in liver functioning affect whole-body metabolism and energy homeostasis.
In this context, adenosine-monophosphate- (AMP-) activated protein kinase (AMPK)
is the downstream component of a protein kinase cascade acting as an
intracellular energy sensor regulating physiological energy dynamics by
limiting anabolic pathways, to prevent excessive adenosine triphosphate (ATP)
utilization, and
by stimulating catabolic processes, to increase ATP production.
Thus, the understanding of the mechanisms by which liver AMPK coordinates hepatic
energy metabolism represents a crucial point of convergence of regulatory signals
monitoring systemic and cellular energy status
Liver AMPK: Structure and Regulation
AMPK, a serine/threonine kinase, is a heterotrimeric complex comprising
a catalytic subunit α and
two regulatory subunits β and γ .
The α subunit has a threonine residue (Thr172) within the activation loop of the kinase
domain, with the C-terminal region being required for association with β and γ subunits.
The β subunit associates with α and γ by means of its C-terminal region , whereas
the γ subunit has four cystathionine β-synthase (CBS) motifs, which
bind AMP or ATP in a competitive manner.
75675.fig.001 (not shown)
Figure 1: Regulation of AMP-activated protein kinase (AMPK) by
(A) direct allosteric activation and
(B) reversible phosphorylation and downstream responses maintaining
intracellular energy balance.
Regulation of liver AMPK activity involves both direct allosteric activation and
reversible phosphorylation. AMPK is allosterically activated by AMP through
binding to the regulatory subunit-γ, which induces a conformational change in
the kinase domain of subunit α that protects AMPK from dephosphorylation
of Thr172, probably by protein phosphatase-2C.
Activation of AMPK requires phosphorylation of Thr172 in its α subunit, which can be
attained by either
(i) tumor suppressor LKB1 kinase following enhancement in the AMP/ATP ratio, a
kinase that plays a crucial role in AMPK-dependent control of liver glucose and
lipid metabolism;
(ii) Ca2+-calmodulin-dependent protein kinase kinase-β (CaMKKβ) that
phosphorylates AMPK in an AMP-independent, Ca2+-dependent manner;
(iii) transforming growth-factor-β-activated kinase-1 (TAK1), an important
kinase in hepatic Toll-like receptor 4 signaling in response to lipopolysaccharide.
Among these kinases, the relevance of CaMKKβ and TAK1 in liver AMPK activation
remains to be established in metabolic stress conditions. Both allosteric and
phosphorylation mechanisms are able to elicit
over 1000-fold increase in AMPK activity, thus allowing
the liver to respond to small changes in energy status in a highly sensitive fashion.
In addition to rapid AMPK regulation through allosterism and reversible phosphorylation
long-term effects of AMPK activation induce changes in hepatic gene expression.
This was demonstrated for
(i) the transcription factor carbohydrate-response element-binding protein (ChREBP),
whose Ser568 phosphorylation by activated AMPK
blocks its DNA binding capacity and glucose-induced gene transcription
under hyperlipidemic conditions;(ii) liver sterol regulatory element-binding
protein-1c (SREBP-1c), whose mRNA and protein expression and those of
its target gene for fatty acid synthase (FAS)
are reduced by metformin-induced AMPK activation,
decreasing lipogenesis and increasing fatty acid oxidation due to
malonyl-CoA depletion;
(iii) transcriptional coactivator transducer of regulated CREB activity-2 (TORC2),
a crucial component of the hepatic gluconeogenic program, was reported
to be phosphorylated by activated AMPK.
This modification leads to subsequent cytoplasmatic sequestration of TORC2 and
inhibition of gluconeogenic gene expression, a mechanism underlying
the plasma glucose-lowering effects of adiponectin and metformin
through AMPK activation by upstream LKB1.
Activation of AMPK in the liver is a key regulatory mechanism controlling glucose
and lipid metabolism,
inhibiting anabolic processes, and
enhancing catabolic pathways in response to different signals, including
energy status,
serum insulin/glucagon ratio,
nutritional stresses,
pharmacological and natural compounds, and
oxidative stress status
Reactive Oxygen Species (ROS) and AMPK Activation
The high energy demands required to cope with all the metabolic functions
of the liver are met by
fatty acid oxidation under conditions of both normal blood glucose levels and
hypoglycemia, whereas
glucose oxidation is favoured in hyperglycemic states, with consequent
generation of ROS.
Under normal conditions, ROS occur at relatively low levels due to their fast processing
by antioxidant mechanisms, whereas at acute or prolonged high ROS levels, severe
oxidation of biomolecules and dysregulation of signal transduction and gene expression
is achieved, with consequent cell death through necrotic and/or apoptotic-signaling
pathways.
Thyroid Hormone (L-3,3′,5-Triiodothyronine, T3), Metabolic Regulation,
and ROS Production
T3 is important for the normal function of most mammalian tissues, with major actions
on O2 consumption and metabolic rate, thus
determining enhancement in fuel consumption for oxidation processes
and ATP repletion.
T3 acts predominantly through nuclear receptors (TR) α and β, forming
functional complexes with retinoic X receptor that
bind to thyroid hormone response elements (TRE) to activate gene expression.
T3 calorigenesis is primarily due to the
induction of enzymes related to mitochondrial electron transport and ATP
synthesis, catabolism, and
some anabolic processes via upregulation of genomic mechanisms.
The net result of T3 action is the enhancement in the rate of O2 consumption of target
tissues such as liver, which may be effected by secondary processes induced by T3
(i) energy expenditure due to higher active cation transport,
(ii) energy loss due to futile cycles coupled to increase in catabolic and anabolic pathways, and
(iii) O2 equivalents used in hepatic ROS generation both in hepatocytes and Kupffer cells
In addition, T3-induced higher rates of mitochondrial oxidative phosphorylation are
likely to induce higher levels of ATP, which are partially balanced by intrinsic uncoupling
afforded by induction of uncoupling proteins by T3. In agreement with this view, the
cytosolic ATP/ADP ratio is decreased in hyperthyroid tissues, due to simultaneous
stimulation of ATP synthesis and consumption.
Regulation of fatty acid oxidation is mainly attained by carnitine palmitoyltransferase Iα (CPT-Iα),
catalyzing the transport of fatty acids from cytosol to mitochondria for β-oxidation,
and acyl-CoA oxidase (ACO),
catalyzing the first rate-limiting reaction of peroxisomal β-oxidation, enzymes that are
induced by both T3 and peroxisome proliferator-activated receptor α (PPAR-α).
Furthermore, PPAR-α-mediated upregulation of CPT-Iα mRNA is enhanced by PPAR-γ
coactivator 1α (PGC-1α), which in turn
augments T3 induction of CPT-Iα expression.
Interestingly, PGC-1α is induced by
T3,
AMPK activation, and
ROS,
thus establishing potential links between
T3 action, ROS generation, and AMPK activation
with the onset of mitochondrial biogenesis and fatty acid β-oxidation.
Liver ROS generation leads to activation of the transcription factors
nuclear factor-κB (NF-κB),
activating protein 1 (AP-1), and
signal transducer and activator of transcription 3 (STAT3)
at the Kupffer cell level, with upregulation of cytokine expression (TNF-α, IL-1, IL-6),
which upon interaction with specific receptors in hepatocytes trigger the expression of
cytoprotective proteins (Figure 3(A)).
These responses and the promotion of hepatocyte and Kupffer-cell proliferation
represent hormetic effects reestablishing
redox homeostasis,
promoting cell survival, and
protecting the liver against ischemia-reperfusion injury.
T3 liver preconditioning also involves the activation of the
Nrf2-Keap1 defense pathway
upregulating antioxidant proteins,
phase-2 detoxifying enzymes, and
multidrug resistance proteins, members of the ATP binding cassette (ABC)
superfamily of transporters (Figure 3(B))
In agreement with T3-induced liver preconditioning, T3 or L-thyroxin afford
preconditioning against IR injury in the heart, in association with
Figure 2: Calorigenic response of thyroid hormone (T3) and its relationship with O2
consumption, reactive oxygen species (ROS) generation, and antioxidant depletion in the liver.
Abbreviations: CYP2E1, cytochrome P450 isoform 2E1; GSH, reduced glutathione; QO2, rate
of O2 consumption; SOD, superoxide dismutase.
475675.fig.003
genomic signaling in T3 calorigenesis and ROS production 475675.fig.003
effects that are achieved through enhanced transcription of TRE-containing genes
Skeletal muscle AMPK activation is characterized by
(i) being a rapid and transient response,
(ii) upstream activation by Ca2+-induced mobilization and CaMKKβ activation,
(iii) upstream upregulation of LKB1 expression, which requires association with STRAD
and MO25 for optimal phosphorylation/activation of AMPK, and
(iv) stimulation of mitochondrial fatty acid β-oxidation.
T3-induced muscle AMPK activation was found to trigger two major downstream
signaling pathways, namely,
(i) peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) mRNA
expression and phosphorylation, a transcriptional regulator for genes related to
mitochondrial biogenesis,
fatty acid oxidation, and
gluconeogenesis and
(ii) cyclic AMP response element binding protein (CREB) phosphorylation, which
in turn induces PGC-1α expression in liver tissue, thus
reinforcing mechanism (i).
These data indicate that AMPK phosphorylation of PGC-1α initiates many of the
important gene regulatory functions of AMPK in skeletal muscle.
In heart, hyperthyroidism increased glycolysis and sarcolemmal GLUT4 levels by the
combined effects of AMPK activation and insulin stimulation, with concomitant increase
in fatty acid oxidation proportional to enhanced cardiac mass and contractile function.
Thyroid Hormone, AMPK Activation, and Liver Preconditioning
Recent studies by our group revealed that administration of a single dose of 0.1 mg T3/kg
to rats activates liver AMPK (Figure 4; unpublished work).
enhancement in phosphorylated AMPK/nonphosphorylated AMPK ratios in T3-
treated rats over control values thatis significant in the time period of 1 to 48
hours after hormone treatment
Administration of a substantially higher dose (0.4 mg T3/kg) resulted in
decreased liver AMPK activation at 4 h to return to control values at 6 h
after treatment
Activation of liver AMPK by T3 may be of relevance in terms of
promotion of fatty acid oxidation for ATP supply,
supporting hepatoprotection against IR injury (Figure 3(C)).
This proposal is based on the high energy demands underlying effective liver
preconditioning for full operation of hepatic
antioxidant, antiapoptotic, and anti-inflammatory mechanisms,
oxidized biomolecules repair or resynthesis,
induction of the homeostatic acute-phase response, and
promotion of hepatocyte and Kupffer cell proliferation,
mechanisms that are needed to cope with the damaging processes set in by IR.
T3 liver preconditioning , in addition to that afforded by
n-3 long-chain polyunsaturated fatty acids given alone or
combined with T3 at lower dosages, or
by iron supplementation,
constitutes protective strategies against hepatic IR injury.
Studies on the molecular mechanisms underlying T3-induced liver AMPK
activation (Figure 4) are currently under assessment in our laboratory.
References
Fernández and L. A. Videla, “Kupffer cell-dependent signaling in thyroid hormone
calorigenesis: possible applications for liver preconditioning,” Current Signal
Transduction Therapy 2009; 4(2): 144–151.
Viollet, B. Guigas, J. Leclerc et al., “AMP-activated protein kinase in the regulation
of hepatic energy metabolism: from physiology to therapeutic perspectives,” Acta
Physiologica 2009; 196(1): 81–98.
Carling, “The AMP-activated protein kinase cascade – A unifying system
for energy control,” Trends in Biochemical Sciences, 2004;. 29(1): 18–24.
E. Kemp, D. Stapleton, D. J. Campbell et al., “AMP-activated protein kinase,
super metabolic regulator,” Biochemical Society Transactions 2003; 31(1):
162–168.
G. Hardie, “AMP-activated protein kinase-an energy sensor that
regulates all ;aspects of cell function,” Genes and Development,
2011; 25(18): 1895–1908.
Woods, P. C. F. Cheung, F. C. Smith et al., “Characterization of AMP-activated
protein kinase βandγ subunits Assembly of the heterotrimeric complex in vitro,”
Journal of Biological Chemistry 1996;271(17): 10282–10290.
Xiao, R. Heath, P. Saiu et al., “Structural basis for AMP binding to mammalian AMP-
activated protein kinase,” Nature 2007; 449(7161): 496–500.
Although adenosine monophosphate activated protein kinase (AMPK) plays a crucial role
in energy metabolism, a direct effect of AMPK modulation on thyroid function has only
recently been reported, and much of its function in the thyroid is currently unknown.
The aim of this study was
to investigate the mechanism of AMPK modulation in iodide uptake.
to investigate the potential of the AMPK inhibitor compound C as an enhancer of
iodide uptake by thyrocytes.
Metformin reduced NIS promoter activity (0.6-fold of control), whereas compound C
stimulated its activity (3.4-fold) after 4 days. This largely coincides with
CRE activation (0.6- and 3.0-fold).
These experiments show that AMPK exerts its effects on iodide uptake, at least partly,
through the CRE element in the NIS promoter. Furthermore, we have used AMPK-alpha1
knockout mice to determine the long-term effects of AMPK inhibition without chemical compounds.
These mice have a less active thyroid, as shown by reduced colloid volume and reduced
responsiveness to thyrotropin.
NIS expression and iodine uptake in thyrocytes
can be modulated by metformin and compound C.
These compounds exert their effect by
modulation of AMPK, which, in turn, regulates
the activation of the CRE element in the NIS promoter.
Overall, this suggests that AMPK modulating compounds may be useful for the
enhancement of iodide uptake by thyrocytes, which could be useful for the
treatment of thyroid cancer patients with radioactive iodine.
AMPK induces a cascade of events within cells in response to the ever changing energy
charge of the cell. The role of AMPK in regulating cellular energy charge places this
enzyme at a central control point in maintaining energy homeostasis.
More recent evidence has shown that AMPK activity can also be regulated by physiological stimuli, independent of the energy charge of the cell, including hormones and nutrients.
Once activated, AMPK-mediated phosphorylation events
How these latter two functions impact obesity and diabetes will be discussed below.
Regulation of AMPK
In the presence of AMP the activity of AMPK is increased approximately 5-fold.
However, more importantly is the role of AMP in regulating the level of phosphorylation
of AMPK. An increased AMP to ATP ratio leads to a conformational change in the γ-subunit
leading to increased phosphorylation and decreased dephosphorylation of AMPK.
The phosphorylation of AMPK results in activation by at least 100-fold. AMPK is
phosphorylated by at least three different upstream AMPK kinases (AMPKKs).
Phosphorylation of AMPK occurs in the α subunit at threonine 172 (T172) which
which phosphorylates and activates AMPK in response to increased calcium.
The distribution of CaMKKβ expression is primarily in the brain with detectable levels
also found in the testes, thymus, and T cells. As described for the Ca2+-mediated
regulation of glycogen metabolism,
increased release of intracellular stores of Ca2+ create a subsequent demand for
ATP.
Activation of AMPK in response to Ca fluxes
provides a mechanism for cells to anticipate the increased demand for ATP.
Evidence has also demonstrated that the serine-threonine kinase, LKB1 (also called
serine-threonine kinase 11, STK11) which is encoded by the Peutz-Jeghers syndrome
tumor suppressor gene, is required for activation of AMPK in response to stress.
The active LKB1 kinase is actually a complex of three proteins:
LKB1,
Ste20-related adaptor (STRAD) and
mouse protein 25 (MO25).
Thus, the enzyme complex is often referred to as LKB1-STRAD-MO25. Phosphorylation
of AMPK by LKB1 also occurs on T172. Unlike the limited distribution of CaMKKβ,
LKB1 is widely expressed, thus making it the primary AMPK-regulating kinase.
Loss of LKB1 activity in adult mouse liver leads to
near complete loss of AMPK activity and
is associated with hyperglycemia.
The hyperglycemia is, in part, due to an increase in the transcription of gluconeogenic
genes. Of particular significance is the increased expression of
the peroxisome proliferator-activated receptor-γ (PPAR-γ) coactivator 1α
(PGC-1α), which drives gluconeogenesis.
Reduction in PGC-1α activity results in normalized blood glucose levels in
LKB1-deficient mice.
The third AMPK phosphorylating kinase is transforming growth factor-β-activated
kinase 1 (TAK1). However, the normal physiological conditions under which TAK1
phosphorylates AMPK are currently unclear.
The effects of AMP are two-fold:
a direct allosteric activation and making AMPK a poorer substrate for
dephosphorylation.
Because AMP affects both
the rate of AMPK phoshorylation in the positive direction and
dephosphorylation in the negative direction,
the cascade is ultrasensitive. This means that
a very small rise in AMP levels can induce a dramatic increase in the activity of
AMPK.
The activity of adenylate kinase, catalyzing the reaction shown below, ensures that
AMPK is highly sensitive to small changes in the intracellular [ATP]/[ADP] ratio.
2 ADP ——> ATP + AMP
Negative allosteric regulation of AMPK also occurs and this effect is exerted by phosphocreatine. As indicated above, the β subunits of AMPK have a glycogen-binding domain, GBD. In muscle, a high glycogen content
represses AMPK activity and
this is likely the result of interaction between the GBD and glycogen,
the GBD of AMPK allows association of the enzyme with the regulation of glycogen metabolism
by placing AMPK in close proximity to one of its substrates glycogen synthase.
AMPK has also been shown to be activated by receptors that are coupled to
phospholipase C-β (PLC-β) and by
hormones secreted by adipose tissue (termed adipokines) such as leptinand adiponectin (discussed below).
Targets of AMPK
The signaling cascades initiated by the activation of AMPK exert effects on
glucose and lipid metabolism,
gene expression and
protein synthesis.
These effects are most important for regulating metabolic events in the liver, skeletal
muscle, heart, adipose tissue, and pancreas.
Demonstration of the central role of AMPK in the regulation of metabolism in response
to events such as nutrient- or exercise-induced stress. Several of the known physiologic
targets for AMPK are included as well as several pathways whose flux is affected by
AMPK activation. Arrows indicate positive effects of AMPK, whereas, T-lines indicate
the resultant inhibitory effects of AMPK action.
The uptake, by skeletal muscle, accounts for >70% of the glucose removal from the
serum in humans. Therefore, it should be obvious that this event is extremely important
for overall glucose homeostasis, keeping in mind, of course, that glucose uptake by
cardiac muscle and adipocytes cannot be excluded from consideration. An important fact
related to skeletal muscle glucose uptake is that this process is markedly impaired in
individuals with type 2 diabetes.
The uptake of glucose increases dramatically in response to stress (such as ischemia) and
exercise and is stimulated by insulin-induced recruitment of glucose transporters
to the plasma membrane, primarily GLUT4. Insulin-independent recruitment of glucose
transporters also occurs in skeletal muscle in response to contraction (exercise).
The activation of AMPK plays an important, albeit not an exclusive, role in the induction of
GLUT4 recruitment to the plasma membrane. The ability of AMPK to stimulate
GLUT4 translocation to the plasma membrane in skeletal muscle is by a different mechanism
than that stimulated by insulin and insulin and AMPK effects are additive.
Under ischemic/hypoxic conditions in the heart the activation of AMPK leads to the
phosphorylation and activation of the kinase activity of phosphofructokinase-2, PFK-2
(6-phosphofructo-2-kinase). The product of the action of PFK-2 (fructose-2,6-bisphosphate,
F2,6BP) is one of the most potent regulators of the rate of flux through glycolysis and gluconeogenesis.
In liver the PKA-mediated phosphorylation of PFK-2 results in conversion of the
enzyme from a kinase that generates F2,6BP to a phosphatase that removes the
2-phosphate thus reducing the levels of the potent allosteric activator of the glycolytic
enzyme 6-phosphfructo-1-kinase, PFK-1 and the potent allosteric inhibitor
of the gluconeogenic enzyme fructose-1,6-bisphosphatase (F1,-6BPase).
It is important to note that like many enzymes, there are multiple isoforms of PFK-2
(at least 4) and neither the liver or the skeletal muscle isoforms contain the AMPK
phosphorylation sites found in the cardiac and inducible (iPFK2) isoforms of PFK-2.
Inducible PFK-2 is expressed in the monocyte/macrophage lineage in response to pro-
inflammatory stimuli. The ability to activate the kinase activity by phosphorylation of
PFK-2 in cardiac tissue and macrophages in response to ischemic conditions allows these
cells to continue to have a source of ATP via anaerobic glycolysis. This phenomenon is
recognized as the Pasteur effect: an increased rate of glycolysis in response to hypoxia.
Of pathological significance is the fact that the inducible form of PFK-2 is commonly
expressed in many tumor cells and this may allow AMPK to play an important role in
protecting tumor cells from hypoxic stress. Indeed, techniques for depleting AMPK in
tumor cells have shown that these cells become sensitized to nutritional stress upon loss
of AMPK activity.
Whereas, stress and exercise are powerful inducers of AMPK activity in skeletal muscle,
additional regulators of its activity have been identified.
Insulin-sensitizing drugs of the thiazolidinedione family (activators of PPAR-γ, see
below) as well as the hypoglycemia drug metformin exert a portion of their effects
through regulation of the activity of AMPK.
As indicated above, the activity of the AMPK activating kinase, LKB1, is critical for
regulating gluconeogenic flux and consequent glucose homeostasis. The action of
metformin in reducing blood glucose levels
requires the activity of LKB1 in the liver for this function.
AMPK-mediated phosphorylation of eNOS leads to increased activity and consequent
NO production and provides a link between metabolic stresses and cardiac function.
In platelets, insulin action leads to an increase in eNOS activity that is
due to its phosphorylation by AMPK.
Activation of NO production in platelets leads to
a decrease in thrombin-induced aggregation, thereby,
limiting the pro-coagulant effects of platelet activation.
The response of platelets to insulin function clearly indicates why disruption in insulin
action is a major contributing factor in the development of the metabolic syndrome
Activation of AMPK leads to a reduction in the level of SREBP
a transcription factor ®ulator of the expression of numerous
lipogenic enzymes
Another transcription factor reduced in response to AMPK activation is
hepatocyte nuclear factor 4α, HNF4α
a member of the steroid/thyroid hormone superfamily.
HNF4α is known to regulate the expression of several liver and
pancreatic β-cell genes such as GLUT2, L-PK and preproinsulin.
Of clinical significance is that mutations in HNF4α are responsible for
maturity-onset diabetes of the young, MODY-1.
Recent evidence indicates that the gene for the carbohydrate-response-element-
binding protein (ChREBP) is a target for AMPK-mediated transcriptional regulation
in the liver. ChREBP is rapidly being recognized as a master regulator of lipid
metabolism in liver, in particular in response to glucose uptake.
The transcription co-activator, p300, is phosphorylated by AMPK
which inhibits interaction of p300 with not only PPARγ but also
the retinoic acid receptor, retinoid X receptor, and
thyroid hormone receptor.
PPARγ is primarily expressed in adipose tissue and thus it was difficult to reconcile how
a drug that was apparently acting only in adipose tissue could lead to improved insulin
sensitivity of other tissues. The answer to this question came when it was discovered that the TZDs stimulated the expression and release of the adipocyte hormone (adipokine),
adiponectin. Adiponectin stimulates glucose uptake and fatty acid oxidation in skeletal
muscle. In addition, adiponectin stimulates fatty acid oxidation in liver while inhibiting
expression of gluconeogenic enzymes in this tissue.
These responses to adiponectin are exerted via activation of AMPK. Another
transcription factor target of AMPK is the forkhead protein, FKHR (now referred to as FoxO1). FoxO1 is involved in the activation of glucose-6-phosphatase expression and,
therefore, loss of FoxO1 activity in response to AMPK activation will lead to reduced
hepatic output of glucose.
This concludes a very complicated perspective that ties together the thyroid hormone
activity, the hypophysis, diabetes mellitus, and AMPK tegulation of metabolism in the
liver, skeletal muscle, adipose tissue, and heart. I also note at this time that there
nongenetic points to be made here:
The tissue specificity of isoenzymes
The modulatory role of AMP:ATP ratio in phosphorylation/dephosphorylation
effects on metabolism tied to AMPK
The tie in of stress or ROS with fast reactions to protect harm to tissues
The relationship of cytokine activation and release to the above metabolic events
The relationship of effective and commonly used diabetes medications to AMPK
mediated processes
The preceding presentation is notable for the importance of proteomic and
metabolomic invetigations in elucidation common chronic and nongenetic diseases
This portion of the discussion is a series of articles on signaling and signaling pathways. Many of the protein-protein interactions or protein-membrane interactions and associated regulatory features have been referred to previously, but the focus of the discussion or points made were different. I considered placing this after the discussion of proteins and how they play out their essential role, but this is quite a suitable place for a progression to what follows. This is introduced by material taken from Wikipedia, which will be followed by a series of mechanisms and examples from the current literature, which give insight into the developments in cell metabolism, with the later goal of separating views introduced by molecular biology and genomics from functional cellular dynamics that are not dependent on the classic view. The work is vast, and this discussion does not attempt to cover it in great depth. It is the first in a series.
Signaling and signaling pathways
Signaling transduction tutorial.
Carbohydrate metabolism
Lipid metabolism
Protein synthesis and degradation
Subcellular structure
Impairments in pathological states: endocrine disorders; stress hypermetabolism; cancer.
Signal transduction occurs when an extracellular signaling[1] molecule activates a specific receptor located on the cell surface or inside the cell. In turn, this receptor triggers a biochemical chain of events inside the cell, creating a response.[2] Depending on the cell, the response alters the cell’s metabolism, shape, gene expression, or ability to divide.[3] The signal can be amplified at any step. Thus, one signaling molecule can cause many responses.[4]
In 1970, Martin Rodbell examined the effects of glucagon on a rat’s liver cell membrane receptor. He noted that guanosine triphosphate disassociated glucagon from this receptor and stimulated the G-protein, which strongly influenced the cell’s metabolism. Thus, he deduced that the G-protein is a transducer that accepts glucagon molecules and affects the cell.[5] For this, he shared the 1994 Nobel Prize in Physiology or Medicine with Alfred G. Gilman.
Signal_transduction_publications_graph
The earliest MEDLINE entry for “signal transduction” dates from 1972.[6] Some early articles used the terms signal transmission and sensory transduction.[7][8] In 2007, a total of 48,377 scientific papers—including 11,211 e review papers—were published on the subject. The term first appeared in a paper’s title in 1979.[9][10] Widespread use of the term has been traced to a 1980 review article by Rodbell:[5][11] Research papers focusing on signal transduction first appeared in large numbers in the late 1980s and early 1990s.[12]
Notch-mediated juxtacrine signal between adjacent cells.
Signal transduction involves the binding of extracellular signaling molecules and ligands to cell-surface receptors that trigger events inside the cell. The combination of messenger with receptor causes a change in the conformation of the receptor, known as receptor activation. This activation is always the initial step (the cause) leading to the cell’s ultimate responses (effect) to the messenger. Despite the myriad of these ultimate responses, they are all directly due to changes in particular cell proteins. Intracellular signaling cascades can be started through cell-substratum interactions; examples are the integrin that binds ligands in the extracellular matrix and steroids.[13] Most steroid hormones have receptors within the cytoplasm and act by stimulating the binding of their receptors to the promoter region of steroid-responsive genes.[14] Examples of signaling molecules include the hormone melatonin,[15] the neurotransmitter acetylcholine[16] and the cytokineinterferon γ.[17]
Signal transduction cascades amplify the signal output
Various environmental stimuli exist that initiate signal transmission processes in multicellular organisms; examples include photons hitting cells in the retina of the eye,[20] and odorants binding to odorant receptors in the nasal epithelium.[21] Certain microbial molecules, such as viral nucleotides and protein antigens, can elicit an immune system response against invading pathogens mediated by signal transduction processes. This may occur independent of signal transduction stimulation by other molecules, as is the case for the toll-like receptor. It may occur with help from stimulatory molecules located at the cell surface of other cells, as with T-cell receptor signaling. Unicellular organisms may respond to environmental stimuli through the activation of signal transduction pathways. For example, slime molds secrete cyclic adenosine monophosphate upon starvation, stimulating individual cells in the immediate environment to aggregate,[22] and yeast cells use mating factors to determine the mating types of other cells and to participate in sexual reproduction.[23] Receptors can be roughly divided into two major classes: intracellular receptors and extracellular receptors.
Extracellular
Extracellular receptors are integral transmembrane proteins and make up most receptors. They span the plasma membrane of the cell, with one part of the receptor on the outside of the cell and the other on the inside. Signal transduction occurs as a result of a ligand binding to the outside; the molecule does not pass through the membrane. This binding stimulates a series of events inside the cell; different types of receptor stimulate different responses and receptors typically respond to only the binding of a specific ligand. Upon binding, the ligand induces a change in the conformation of the inside part of the receptor.[24] These result in either the activation of an enzyme in the receptor or the exposure of a binding site for other intracellular signaling proteins within the cell, eventually propagating the signal through the cytoplasm.
In eukaryotic cells, most intracellular proteins activated by a ligand/receptor interaction possess an enzymatic activity; examples include tyrosine kinase and phosphatases. Some of them create second messengers such as cyclic AMP and IP3, the latter controlling the release of intracellular calcium stores into the cytoplasm. Other activated proteins interact with adaptor proteins that facilitate signalling protein interactions and coordination of signalling complexes necessary to respond to a particular stimulus. Enzymes and adaptor proteins are both responsive to various second messenger molecules.
Many adaptor proteins and enzymes activated as part of signal transduction possess specialized protein domains that bind to specific secondary messenger molecules. For example, calcium ions bind to the EF hand domains of calmodulin, allowing it to bind and activate calmodulin-dependent kinase. PIP3 and other phosphoinositides do the same thing to the Pleckstrin homology domains of proteins such as the kinase protein AKT.
G protein-coupled
G protein-coupled receptors (GPCRs) are a family of integral transmembrane proteins that possess seven transmembrane domains and are linked to a heterotrimeric G protein. Many receptors are in this family, including adrenergic receptors and chemokine receptors.
Arrestin binding to active GPCR kinase (GRK)-phosphorylated GPCRs blocks G protein coupling
Signal transduction by a GPCR begins with an inactive G protein coupled to the receptor; it exists as a heterotrimer consisting of Gα, Gβ, and Gγ.[25] Once the GPCR recognizes a ligand, the conformation of the receptor changes to activate the G protein, causing Gα to bind a molecule of GTP and dissociate from the other two G-protein subunits. The dissociation exposes sites on the subunits that can interact with other molecules.[26] The activated G protein subunits detach from the receptor and initiate signaling from many downstream effector proteins such as phospholipases and ion channels, the latter permitting the release of second messenger molecules.[27] The total strength of signal amplification by a GPCR is determined by the lifetimes of the ligand-receptor complex and receptor-effector protein complex and the deactivation time of the activated receptor and effectors through intrinsic enzymatic activity.
A study was conducted where a point mutation was inserted into the gene encoding the chemokine receptor CXCR2; mutated cells underwent a malignant transformation due to the expression of CXCR2 in an active conformation despite the absence of chemokine-binding. This meant that chemokine receptors can contribute to cancer development.[28]
Tyrosine and histidine kinase
Receptor tyrosine kinases (RTKs) are transmembrane proteins with an intracellular kinase domain and an extracellular domain that binds ligands; examples include growth factor receptors such as the insulin receptor.[29] To perform signal transduction, RTKs need to form dimers in the plasma membrane;[30] the dimer is stabilized by ligands binding to the receptor. The interaction between the cytoplasmic domains stimulates the autophosphorylation of tyrosines within the domains of the RTKs, causing conformational changes. Subsequent to this, the receptors’ kinase domains are activated, initiating phosphorylation signaling cascades of downstream cytoplasmic molecules that facilitate various cellular processes such as cell differentiation and metabolism.[29]
As is the case with GPCRs, proteins that bind GTP play a major role in signal transduction from the activated RTK into the cell. In this case, the G proteins are members of the Ras, Rho, and Raf families, referred to collectively as small G proteins. They act as molecular switches usually tethered to membranes by isoprenyl groups linked to their carboxyl ends. Upon activation, they assign proteins to specific membrane subdomains where they participate in signaling. Activated RTKs in turn activate small G proteins that activate guanine nucleotide exchange factors such as SOS1. Once activated, these exchange factors can activate more small G proteins, thus amplifying the receptor’s initial signal. The mutation of certain RTK genes, as with that of GPCRs, can result in the expression of receptors that exist in a constitutively activate state; such mutated genes may act as oncogenes.[31]
Histidine-specific protein kinases are structurally distinct from other protein kinases and are found in prokaryotes, fungi, and plants as part of a two-component signal transduction mechanism: a phosphate group from ATP is first added to a histidine residue within the kinase, then transferred to an aspartate residue on a receiver domain on a different protein or the kinase itself, thus activating the aspartate residue.[32]
Integrin
integrin-mediated signal transduction
An overview of integrin-mediated signal transduction, adapted from Hehlgens et al. (2007).[33]
Integrins are produced by a wide variety of cells; they play a role in cell attachment to other cells and the extracellular matrix and in the transduction of signals from extracellular matrix components such as fibronectin and collagen. Ligand binding to the extracellular domain of integrins changes the protein’s conformation, clustering it at the cell membrane to initiate signal transduction. Integrins lack kinase activity; hence, integrin-mediated signal transduction is achieved through a variety of intracellular protein kinases and adaptor molecules, the main coordinator being integrin-linked kinase.[33] As shown in the picture to the right, cooperative integrin-RTK signalling determines the timing of cellular survival, apoptosis, proliferation, and differentiation.
Important differences exist between integrin-signalling in circulating blood cells and non-circulating cells such as epithelial cells; integrins of circulating cells are normally inactive. For example, cell membrane integrins on circulating leukocytes are maintained in an inactive state to avoid epithelial cell attachment; they are activated only in response to stimuli such as those received at the site of an inflammatory response. In a similar manner, integrins at the cell membrane of circulating platelets are normally kept inactive to avoid thrombosis. Epithelial cells (which are non-circulating) normally have active integrins at their cell membrane, helping maintain their stable adhesion to underlying stromal cells that provide signals to maintain normal functioning.[34]
Toll gate
When activated, toll-like receptors (TLRs) take adapter molecules within the cytoplasm of cells in order to propagate a signal. Four adaptor molecules are known to be involved in signaling, which are Myd88, TIRAP, TRIF, and TRAM.[35][36][37] These adapters activate other intracellular molecules such as IRAK1, IRAK4, TBK1[disambiguation needed], and IKKi that amplify the signal, eventually leading to the induction or suppression of genes that cause certain responses. Thousands of genes are activated by TLR signaling, implying that this method constitutes an important gateway for gene modulation.
Ligand-gated ion channel
A ligand-gated ion channel, upon binding with a ligand, changes conformation to open a channel in the cell membrane through which ions relaying signals can pass. An example of this mechanism is found in the receiving cell of a neural synapse. The influx of ions that occurs in response to the opening of these channels induces action potentials, such as those that travel along nerves, by depolarizing the membrane of post-synaptic cells, resulting in the opening of voltage-gated ion channels.
An example of an ion allowed into the cell during a ligand-gated ion channel opening is Ca2+; it acts as a second messenger initiating signal transduction cascades and altering the physiology of the responding cell. This results in amplification of the synapse response between synaptic cells by remodelling the dendritic spines involved in the synapse.
Ion transporters and channels in mammalian choroidal epithelium
Intracellular
Extracellular receptors are integral transmembrane proteins and make up most receptors. They span the plasma membrane of the cell, with one part of the receptor on the outside of the cell and the other on the inside. Signal transduction occurs as a result of a ligand binding to the outside; the molecule does not pass through the membrane. This binding stimulates a series of events inside the cell; different types of receptor stimulate different responses and receptors typically respond to only the binding of a specific ligand. Upon binding, the ligand induces a change in the conformation of the inside part of the receptor.[24] These result in either the activation of an enzyme in the receptor or the exposure of a binding site for other intracellular signaling proteins within the cell, eventually propagating the signal through the cytoplasm.
Understanding these receptors and identifying their ligands and the resulting signal transduction pathways represent a major conceptual advance
intercellular signaling
conformational-rearrangements
membrane protein receptor binds with hormone
The multiple protein-dependent steps in signal transduction
In eukaryotic cells, most intracellular proteins activated by a ligand/receptor interaction possess an enzymatic activity; examples include tyrosine kinase and phosphatases. Some of them create second messengers such as cyclic AMP and IP3, the latter controlling the release of intracellular calcium stores into the cytoplasm. Other activated proteins interact with adaptor proteins that facilitate signalling protein interactions and coordination of signalling complexes necessary to respond to a particular stimulus. Enzymes and adaptor proteins are both responsive to various second messenger molecules.
Ca++ exchange
Many adaptor proteins and enzymes activated as part of signal transduction possess specialized protein domains that bind to specific secondary messenger molecules. For example, calcium ions bind to the EF hand domains of calmodulin, allowing it to bind and activate calmodulin-dependent kinase. PIP3 and other phosphoinositides do the same thing to the Pleckstrin homology domains of proteins such as the kinase protein AKT.
G protein-coupled
G protein-coupled receptors (GPCRs) are a family of integral transmembrane proteins that possess seven transmembrane domains and are linked to a heterotrimeric G protein. Many receptors are in this family, including adrenergic receptors and chemokine receptors.
membrane_receptor_g protein
intracellular_receptor_steroid
Signal transduction by a GPCR begins with an inactive G protein coupled to the receptor; it exists as a heterotrimer consisting of Gα, Gβ, and Gγ.[25] Once the GPCR recognizes a ligand, the conformation of the receptor changes to activate the G protein, causing Gα to bind a molecule of GTP and dissociate from the other two G-protein subunits. The dissociation exposes sites on the subunits that can interact with other molecules.[26] The activated G protein subunits detach from the receptor and initiate signaling from many downstream effector proteins such as phospholipases and ion channels, the latter permitting the release of second messenger molecules.[27] The total strength of signal amplification by a GPCR is determined by the lifetimes of the ligand-receptor complex and receptor-effector protein complex and the deactivation time of the activated receptor and effectors through intrinsic enzymatic activity.
A study was conducted where a point mutation was inserted into the gene encoding the chemokine receptor CXCR2; mutated cells underwent a malignant transformation due to the expression of CXCR2 in an active conformation despite the absence of chemokine-binding. This meant that chemokine receptors can contribute to cancer development.[28]
Tyrosine and histidine kinase
Receptor tyrosine kinases (RTKs) are transmembrane proteins with an intracellular kinase domain and an extracellular domain that binds ligands; examples include growth factor receptors such as the insulin receptor.[29] To perform signal transduction, RTKs need to form dimers in the plasma membrane;[30] the dimer is stabilized by ligands binding to the receptor. The interaction between the cytoplasmic domains stimulates the autophosphorylation of tyrosines within the domains of the RTKs, causing conformational changes. Subsequent to this, the receptors’ kinase domains are activated, initiating phosphorylation signaling cascades of downstream cytoplasmic molecules that facilitate various cellular processes such as cell differentiation and metabolism.[29]
As is the case with GPCRs, proteins that bind GTP play a major role in signal transduction from the activated RTK into the cell. In this case, the G proteins are members of the Ras, Rho, and Raf families, referred to collectively as small G proteins. They act as molecular switches usually tethered to membranes by isoprenyl groups linked to their carboxyl ends. Upon activation, they assign proteins to specific membrane subdomains where they participate in signaling. Activated RTKs in turn activate small G proteins that activate guanine nucleotide exchange factors such as SOS1. Once activated, these exchange factors can activate more small G proteins, thus amplifying the receptor’s initial signal. The mutation of certain RTK genes, as with that of GPCRs, can result in the expression of receptors that exist in a constitutively activate state; such mutated genes may act as oncogenes.[31]
Histidine-specific protein kinases are structurally distinct from other protein kinases and are found in prokaryotes, fungi, and plants as part of a two-component signal transduction mechanism: a phosphate group from ATP is first added to a histidine residue within the kinase, then transferred to an aspartate residue on a receiver domain on a different protein or the kinase itself, thus activating the aspartate residue.[32]
Integrin
integrin-mediated signal transduction
An overview of integrin-mediated signal transduction, adapted from Hehlgens et al. (2007).[33]
Integrins are produced by a wide variety of cells; they play a role in cell attachment to other cells and the extracellular matrix and in the transduction of signals from extracellular matrix components such as fibronectin and collagen. Ligand binding to the extracellular domain of integrins changes the protein’s conformation, clustering it at the cell membrane to initiate signal transduction. Integrins lack kinase activity; hence, integrin-mediated signal transduction is achieved through a variety of intracellular protein kinases and adaptor molecules, the main coordinator being integrin-linked kinase.[33] As shown in the picture to the right, cooperative integrin-RTK signalling determines the timing of cellular survival, apoptosis, proliferation, and differentiation.
Platelet signaling pathways
Protein ubiquitylation
ubiquitylation-is-a-multistep-reaction.
Important differences exist between integrin-signaling in circulating blood cells and non-circulating cells such as epithelial cells; integrins of circulating cells are normally inactive. For example, cell membrane integrins on circulating leukocytes are maintained in an inactive state to avoid epithelial cell attachment; they are activated only in response to stimuli such as those received at the site of an inflammatory response. In a similar manner, integrins at the cell membrane of circulating platelets are normally kept inactive to avoid thrombosis. Epithelial cells (which are non-circulating) normally have active integrins at their cell membrane, helping maintain their stable adhesion to underlying stromal cells that provide signals to maintain normal functioning.[34]
Toll gate
When activated, toll-like receptors (TLRs) take adapter molecules within the cytoplasm of cells in order to propagate a signal. Four adaptor molecules are known to be involved in signaling, which are Myd88, TIRAP, TRIF, and TRAM.[35][36][37] These adapters activate other intracellular molecules such as IRAK1, IRAK4, TBK1[disambiguation needed], and IKKi that amplify the signal, eventually leading to the induction or suppression of genes that cause certain responses. Thousands of genes are activated by TLR signaling, implying that this method constitutes an important gateway for gene modulation.
SignalTrans
Ligand-gated ion channel
A ligand-gated ion channel, upon binding with a ligand, changes conformation to open a channel in the cell membrane through which ions relaying signals can pass. An example of this mechanism is found in the receiving cell of a neural synapse. The influx of ions that occurs in response to the opening of these channels induces action potentials, such as those that travel along nerves, by depolarizing the membrane of post-synaptic cells, resulting in the opening of voltage-gated ion channels.
An example of an ion allowed into the cell during a ligand-gated ion channel opening is Ca2+; it acts as a second messenger initiating signal transduction cascades and altering the physiology of the responding cell. This results in amplification of the synapse response between synaptic cells by remodelling the dendritic spines involved in the synapse.
Ion transporters and channels in mammalian choroidal epithelium
Intracellular
Intracellular receptors, such as nuclear receptors and cytoplasmic receptors, are soluble proteins localized within their respective areas. The typical ligands for nuclear receptors are lipophilic hormones like the steroid hormones testosterone and progesterone and derivatives of vitamins A and D. To initiate signal transduction, the ligand must pass through the plasma membrane by passive diffusion. On binding with the receptor, the ligands pass through the nuclear membrane into the nucleus, enabling gene transcription and protein production.
Signal Transduction
Activated nuclear receptors attach to the DNA at receptor-specific hormone-responsive element (HRE) sequences, located in the promoter region of the genes activated by the hormone-receptor complex. Due to their enabling gene transcription, they are alternatively called inductors of gene expression. All hormones that act by regulation of gene expression have two consequences in their mechanism of action; their effects are produced after a characteristically long period of time and their effects persist for another long period of time, even after their concentration has been reduced to zero, due to a relatively slow turnover of most enzymes and proteins that would either deactivate or terminate ligand binding onto the receptor.
Signal transduction via these receptors involves little proteins, but the details of gene regulation by this method are not well-understood. Nucleic receptors have DNA-binding domains containing zinc fingers and a ligand-binding domain; the zinc fingers stabilize DNA binding by holding its phosphate backbone. DNA sequences that match the receptor are usually hexameric repeats of any kind; the sequences are similar but their orientation and distance differentiate them. The ligand-binding domain is additionally responsible for dimerization of nucleic receptors prior to binding and providing structures for transactivation used for communication with the translational apparatus.
signal-transduction-in-protease-signaling-
protein changes in biological mechanisms
Steroid receptors are a subclass of nuclear receptors located primarily within the cytosol; in the absence of steroids, they cling together in an aporeceptor complex containing chaperone or heatshock proteins (HSPs). The HSPs are necessary to activate the receptor by assisting the protein to fold in a way such that the signal sequence enabling its passage into the nucleus is accessible. Steroid receptors, on the other hand, may be repressive on gene expression when their transactivation domain is hidden; activity can be enhanced by phosphorylation of serine residues at their N-terminal as a result of another signal transduction pathway, a process called crosstalk.
Structure of the N-terminal domain of the yeast Hsp90 chaperone
Pincer movement of Hsp90 coupled to the ATPase cycle. NTD = N-terminal domain, MD = middle domain, CTD = C-terminal domain.
Retinoic acid receptors are another subset of nuclear receptors. They can be activated by an endocrine-synthesized ligand that entered the cell by diffusion, a ligand synthesised from a precursor like retinol brought to the cell through the bloodstream or a completely intracellularly synthesised ligand like prostaglandin. These receptors are located in the nucleus and are not accompanied by HSPs; they repress their gene by binding to their specific DNA sequence when no ligand binds to them, and vice versa.
Certain intracellular receptors of the immune system are cytoplasmic receptors; recently identified NOD-like receptors (NLRs) reside in the cytoplasm of some eukaryotic cells and interact with ligands using a leucine-rich repeat (LRR) motif similar to TLRs. Some of these molecules like NOD2 interact with RIP2 kinase that activates NF-κB signaling, whereas others like NALP3 interact with inflammatory caspases and initiate processing of particular cytokines like interleukin-1β.[38][39]
Cell signaling
signaling pathjways map
Cell signalling is part of a complex system of communication that governs basic cellular activities and coordinates cell actions. The ability of cells to perceive and correctly respond to their microenvironment is the basis of development, tissue repair, and immunity as well as normal tissue homeostasis. Errors in cellular information processing are responsible for diseases such as cancer, autoimmunity, and diabetes. By understanding cell signalling, diseases may be treated effectively and, theoretically, artificial tissues may be created.
Traditional work in biology has focused on studying individual parts of cell signaling pathways. Systems biology research helps us to understand the underlying structure of cell signaling networks and how changes in these networks may affect the transmission and flow of information. Such networks are complex systems in their organization and may exhibit a number of emergent properties. Long-range allostery is often a significant component of cell signaling events.[1]
Enzyme_Model allosterism
Classification
Signaling within, between, and among cells is subdivided into the following classifications:
Intracrine signals are produced by the target cell that stay within the target cell.
Autocrine signals are produced by the target cell, are secreted, and effect the target cell itself via receptors. Sometimes autocrine cells can target cells close by if they are the same type of cell as the emitting cell. An example of this are immune cells.
Juxtacrine signals target adjacent (touching) cells. These signals are transmitted along cell membranes via protein or lipid components integral to the membrane and are capable of affecting either the emitting cell or cells immediately adjacent.
Paracrine signals target cells in the vicinity of the emitting cell. Neurotransmitters represent an example.
Endocrine signals target distant cells. Endocrine cells produce hormones that travel through the blood to reach all parts of the body.
Notch-mediated juxtacrine signal between adjacent cells.
Notch-mediated juxtacrine signal between adjacent cells.
Some cell–cell communication requires direct cell–cell contact. Some cells can form gap junctions that connect their cytoplasm to the cytoplasm of adjacent cells. In cardiac muscle, gap junctions between adjacent cells allows for action potential propagation from the cardiac pacemaker region of the heart to spread and coordinately cause contraction of the heart.
The notch signaling mechanism is an example of juxtacrine signaling (also known as contact-dependent signaling) in which two adjacent cells must make physical contact in order to communicate. This requirement for direct contact allows for very precise control of cell differentiation during embryonic development. In the worm Caenorhabditis elegans, two cells of the developing gonad each have an equal chance of terminally differentiating or becoming a uterine precursor cell that continues to divide. The choice of which cell continues to divide is controlled by competition of cell surface signals. One cell will happen to produce more of a cell surface protein that activates the Notch receptor on the adjacent cell. This activates a feedback loop or system that reduces Notch expression in the cell that will differentiate and that increases Notch on the surface of the cell that continues as a stem cell.[5]
Many cell signals are carried by molecules that are released by one cell and move to make contact with another cell. Endocrine signals are called hormones. Hormones are produced by endocrine cells and they travel through the blood to reach all parts of the body. Specificity of signaling can be controlled if only some cells can respond to a particular hormone. Paracrine signals such as retinoic acid target only cells in the vicinity of the emitting cell.[6]Neurotransmitters represent another example of a paracrine signal. Some signaling molecules can function as both a hormone and a neurotransmitter. For example, epinephrine and norepinephrine can function as hormones when released from the adrenal gland and are transported to the heart by way of the blood stream. Norepinephrine can also be produced by neurons to function as a neurotransmitter within the brain.[7]Estrogen can be released by the ovary and function as a hormone or act locally via paracrine or autocrine signaling.[8] Active species of oxygen and nitric oxide can also act as cellular messengers. This process is dubbed redox signaling.
Signaling Pathways
Cell Signaling Biology
Michael J. Berridge
Module 2
Cell Signaling Pathways
The nine membrane-bound adenylyl cyclases (AC1–AC9) have a similar domain structure. The single polypeptide has a tandem repeat of six transmembrane domains (TM) with TM1- -TM6 in one repeat and TM7- -TM12 in the other. Each TM cassette is followed by large cytoplasmic domains (C1 and C2), which contain the catalytic regions that convert ATP into cyclic AMP. As shown in the lower panel, the C1 and C2 domains come together to form a heterodimer. The ATP-binding site is located at the interface between these two domains. The soluble AC10 isoform lacks the transmembrane regions, but it retains the C1 and C2 domains that are responsible for catalysis www.cellsignallingbiology.orghttp://www.biochemj.org/csb/002/csb002.pdf
Resources:
Elucidate Target-Specific Pathways With a Suite of Cellular Assays
DiscoveRx® offers a comprehensive collection of cell-based pathway indicator assays designed to detect activation or inhibition of complex signal transduction pathways in response to compound treatment. Based on the proven PathHunter® technology, These biosensor cell lines allow you to measure distinct events within a variety of pathways involved in compound toxicity, cholesterol metabolism, antioxidant function, DNA damage and ER stress. In combination with our biosensor cell lines with fast and simple chemiluminescent detection, DiscoveRx Pathway Signaling assays will help you generate cellular pathway selectivity profiles of your compounds without relying on reporter gene assays or complex phenotypic screens. – See more at: http://www.discoverx.com/targets/signaling-pathways?gclid=CPPrxrrli8ACFSdp7AodO2IADQ#sthash.OhK3iKl4.dpuf
DiscoveRx® offers a comprehensive collection of cell-based pathway indicator assays designed to detect activation or inhibition of complex signal transduction pathways in response to compound treatment. Based on the proven PathHunter® technology, These biosensor cell lines allow you to measure distinct events within a variety of pathways involved in compound toxicity, cholesterol metabolism, antioxidant function, DNA damage and ER stress. – See more at: http://www.discoverx.com/targets/signaling-pathways#sthash.ZTb5UXVO.dpuf
inhibitors of signal transduction pathway
Inhibitors of MAPK Signaling Pathway
jak-stat
Nrf2 signaling in ARE-mediated coordinated activation of defensive genes
Regulation of AMPK
metabolic pathways
On these resource pages you can find signaling pathway diagrams, research overviews, relevant antibody products, publications, and other research resources organized by topic. The pathway diagrams associated with these topics have been assembled by CST scientists and outside experts to provide succinct and current overviews of selected signaling pathways. Please send suggestions for developing new pathways to info@cellsignal.com. Protein nodes in each pathway diagram are linked to specific antibody product information or, optionally, to protein-specific listings in the PhosphoSitePlus® database of post-translational modifications.
The serine/threonine kinase Akt/PKB exists as three isoforms in mammals. Akt1 has a wide tissue distribution, whereas Akt2 is found predominantly in muscle and fat cells and Akt3 is expressed in testes and brain. Akt regulates multiple biological processes including cell survival, proliferation, growth, and glycogen metabolism. Various growth factors, hormones, and cytokines activate Akt by binding their cognate receptor tyrosine kinase (RTK), cytokine receptor, or GPCR and triggering activation of the lipid kinase PI3K, which generates PIP3 at the plasma membrane. Akt binds PIP3 through its pleckstrin homology (PH) domain, resulting in translocation of Akt to the membrane. Akt is activated through a dual phosphorylation mechanism. PDK1, which is also brought to the membrane through its PH domain, phosphorylates Akt within its activation loop at Thr308. A second phosphorylation at Ser473 within the carboxy terminus is also required for activity and is carried out by the mTOR-rictor complex, mTORC2.
PTEN, a lipid phosphatase that catalyzes the dephosphorylation of PIP3, is a major negative regulator of Akt signaling. Loss of PTEN function has been implicated in many human cancers. Akt activity is also negatively regulated by the phosphatases PP2A and PHLPP, as well as by the chemical modulators wortmannin and LY294002, both of which are inhibitors of PI3K.
Activated Akt phosphorylates a large number of downstream substrates containing the consensus sequence RXRXXS/T. One of its primary functions is to promote cell growth and protein synthesis through regulation of the mTOR signaling pathway. Akt directly phosphorylates and activates mTOR, as well as inhibits the mTOR inhibitor proteins PRAS40 and tuberin (TSC2). Combined, these actions promote cell growth and G1 cell cycle progression through signaling via p70 S6 Kinase and inhibition of 4E-BP1.
Phosphofructokinase mechanism
GSK-3 is a primary target of Akt and inhibitory phosphorylation of GSK-3α (Ser21) or GSK-3β (Ser9) has numerous cellular effects such as promoting glycogen metabolism, cell cycle progression, regulation of wnt signaling, and formation of neurofibrillary tangles in Alzheimers disease. Akt promotes cell survival directly by its ability to phosphorylate and inactivate several pro-apoptotic targets, including Bad, Bim, Bax, and the forkhead (FoxO1/3a) transcription factors. Akt also plays an important role in metabolism and insulin signaling. Insulin receptor signaling through Akt promotes Glut4 translocation through activation of AS160 and TBC1D1, resulting in increased glucose uptake. Akt regulates glycolysis through phosphorylation of PFK and hexokinase, and plays a significant role in aerobic glycolysis of cancer cells, also known as the Warburg Effect.
Aberrant Akt signaling is the underlying defect found in several pathologies. Akt is one of the most frequently activated kinases in human cancer as constitutively active Akt can promote unregulated cell proliferation. Abnormalities in Akt2 signaling can result in diabetes due to defects in glucose homeostasis. Akt is also a key player in cardiovascular disease through its role in cardiac growth, angiogenesis, and hypertrophy.
The mammalian target of rapamycin (mTOR) is an atypical serine/threonine kinase that is present in two distinct complexes. mTOR complex 1 (mTORC1) is composed of mTOR, Raptor, GβL (mLST8), and Deptor and is partially inhibited by rapamycin. mTORC1 integrates multiple signals reflecting the availability of growth factors, nutrients, or energy to promote either cellular growth when conditions are favorable or catabolic processes during stress or when conditions are unfavorable. Growth factors and hormones (e.g. insulin) signal to mTORC1 via Akt, which inactivates TSC2 to prevent inhibition of mTORC1. Alternatively, low ATP levels lead to the AMPK-dependent activation of TSC2 and phosphorylation of raptor to reduce mTORC1 signaling. Amino acid availability is signaled to mTORC1 via a pathway involving the Rag and Ragulator (LAMTOR1-3) proteins. Active mTORC1 has a number of downstream biological effects including translation of mRNA via the phosphorylation of downstream targets (4E-BP1 and p70 S6 Kinase), suppression of autophagy (Atg13, ULK1), ribosome biogenesis, and activation of transcription leading to mitochondrial metabolism or adipogenesis. The mTOR complex 2 (mTORC2) is composed of mTOR, Rictor, GβL, Sin1, PRR5/Protor-1, and Deptor and promotes cellular survival by activating Akt. mTORC2 also regulates cytoskeletal dynamics by activating PKCα and regulates ion transport and growth via SGK1 phosphorylation. Aberrant mTOR signaling is involved in many disease states including cancer, cardiovascular disease, and metabolic disorders.
We would like to thank Carson Thoreen and Prof. David Sabatini, Whitehead Institute for Biomedical Research, MIT, Cambridge, MA, for reviewing this diagram. revised November 2012
Protein Folding
conformational-rearrangements
Pincer movement of Hsp90 coupled to the ATPase cycle. NTD = N-terminal domain, MD = middle domain, CTD = C-terminal domain.
Heat Shock Proteins (HSPs) form seven families (small HSPs (sHSPs), HSP10, 40, 60, 70, 90, and 100) of molecular chaperone proteins that play a central role in the cellular resistance to stress and actin organization. They are involved in the proper folding of proteins and the recognition and refolding of misfolded proteins. HSP expression is induced by a variety of environmental stresses, including heat, hypoxia, nutrient deficiency, free radicals, toxins, ischemia, and UV radiation. HSP27 is a member of the sHSP family. It is phosphorylated at Ser15, Ser78, and Ser82 by MAPKAPK-2 as a result of the activation of the p38 MAP kinase pathway. Phosphorylation and increased concentration of HSP27 has been implicated in actin polymerization and reorganization. HSP70 and HSP90 interact with unfolded proteins to prevent irreversible aggregation and catalyze the refolding of their substrates in an ATP- and co-chaperone-dependent manner. HSP70 has a broad range of substrates including newly synthesized and denatured proteins, while HSP90 tends to have a more limited subset of substrates, most of which are signaling molecules. HSP70 and HSP90 are also essential for the maturation and inactivation of nuclear hormones and other signaling molecules.
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