Posts Tagged ‘Pseudogene’

Long Noncoding RNA Network regulates PTEN Transcription

Posted in Genome Biology, Genomic Testing: Methodology for Diagnosis, International Global Work in Pharmaceutical, Molecular Genetics & Pharmaceutical, tagged Brain tumor, breast cancer, DNA, Karolinska Institutet, Long non-coding RNA, Per Johnsson, Pseudogene, PTEN, RNA, Scripps Research Institute, Tumor suppressor gene on March 6, 2013| 2 Comments »

Long Noncoding RNA Network regulates PTEN Transcription

Author: Larry H Bernstein, MD, FCAP

Scientists Find Surprising New Influence On Cancer Genes

A pseudogene long noncoding RNA networkregulates PTEN transcription and translation in human cells
Per Johnsson, A Ackley, L Vidarsdottir, Weng-Onn Lui, M Corcoran, D Grandér, and KV Morris

a new study led by scientists at The Scripps Research Institute (TSRI) shows how

pseudogenes can regulate the activity of a cancer-related gene called PTEN.

The study also shows that pseudogenes can be targeted to control PTEN’s activity.

http://Nature_StructuralMolecularBiology.com/pseudogene_long_noncoding_RNA_network_regulates_PTEN-in_human_cells/KV_Morris/

Mol Cancer. 2011; 10: 38. Published online 2011 April 13. doi: 10.1186/1476-4598-10-38 PMCID: PMC3098824

New Type of Gene That Regulates Tumour Suppressor PTEN Identified

Feb. 24, 2013 — Researchers at Karolinska Institutet in Sweden have identified a new so-called pseudogene that regulates the tumour-suppressing PTEN gene.
They hope that this pseudogene will be able to control PTEN to

reverse the tumour process,
make the cancer tumour more sensitive to chemotherapy and
prevent the development of resistance.

Health & Medicine

Reference

The findings, which are published in the scientific journal Nature Structural and Molecular Biology, can be of significance in

the future development of cancer drugs.

The development of tumours coincides with the activation of several cancer genes as well as the inactivation of other tumour-suppressing genes owing to

damage to the DNA and
to the fact that

the cancer cells manage to switch off the transcription of tumour-suppressor genes.

To identify what might be regulating this silencing, the researchers studied PTEN,

one of the most commonly inactivated tumour-suppressor genes.

It has long been believed that the switching-off process is irreversible, but the team has now shown that

silenced PTEN genes in tumour cells can be ‘rescued’ and
re-activated by a ‘pseudogene’,
- a type of gene that, unlike normal genes,
- does not encode an entire protein.

“We identified a new non-protein encoding pseudogene, which

determines whether the expression of PTEN
- is to be switched on or off,”

says research team member Per Johnsson, at Karolinska Institutet’s Department of Oncology-Pathology. “What makes this case spectacular is that the gene

only produces RNA,
the protein’s template.

It is this RNA that, through a sequence of mechanisms,

regulates PTEN.

Pseudogenes have been known about for many years, but

it was thought that they were only junk material.”

No less than 98 per cent of human DNA consists of non-protein encoding genes (i.e. pseudogenes), and by studying these formerly neglected genes the researchers

have begun to understand that they are very important and
- can have an effect without encoding proteins.

Using model systems, the team has shown that the new pseudogene can

control the expression of PTEN and
- make tumours more responsive to conventional chemotherapy.

Pre Johnssom suggests “we might one day be able to re-programme cancer cells

to proliferate less,
become more normal, and that
resistance to chemotherapy can hopefully be avoided.

“We also believe that our findings can be very important for the future development of cancer drugs. The human genome conceals no less than 15,000 or so pseudogenes, and it’s not unreasonable to think

that many of them are relevant to diseases such as cancer.”

The study was conducted in collaboration with scientists at The Scripps Research Institute, USA, and the University of New South Wales, Australia, and was made possible with

grants from the Swedish Childhood Cancer Foundation, the Swedish Cancer Society, the Cancer Research Funds of Radiumhemmet, Karolinska Institutet’s KID programme for doctoral studies, the Swedish Research Council, the Erik and Edith Fernström Foundation for Medical Research, the National Institute of Allergy and Infectious Diseases, the National Cancer Institute and the National Institutes of Health.

The functional role of long non-coding RNA in human carcinomas
EA Gibb, CJ Brown, and WL Lam
Long non-coding RNAs (lncRNAs) are emerging as new players in the cancer paradigm demonstrating potential roles in both oncogenic and tumor suppressive pathways. These novel genes are frequently

aberrantly expressed in a variety of human cancers,

however the biological functions of the vast majority remain unknown. Recently, evidence has begun to accumulate describing the molecular mechanisms by which these RNA species function, providing insight into

the functional roles they may play in tumorigenesis.

In this review, we highlight the emerging functional role of lncRNAs in human cancer.

One of modern biology’s great surprises was the discovery that the human genome encodes only ~20,000 protein-coding genes, representing <2% of the total genome sequence [1,2]. However, with the advent of

tiling resolution genomic microarrays and
whole genome and transcriptome sequencing technologies
- it was determined that at least 90% of the genome is actively transcribed [3,4].

The human transcriptome was found to be more complex than

a collection of protein-coding genes and their splice variants; showing
- extensive antisense,
- overlapping and non-coding RNA (ncRNA) expression [5-10].

Although initially argued to be spurious transcriptional noise, recent evidence suggests that the proverbial “dark matter” of the genome

may play a major biological role in cellular development and metabolism [11-17].

One such player, the newly discovered long non-coding RNA (lncRNA) genes, demonstrate

developmental and tissue specific expression patterns, and
aberrant regulation in a variety of diseases, including cancer [18-27].

NcRNAs are loosely grouped into two major classes based on transcript size; small ncRNAs and lncRNAs [28-30].

Small ncRNAs are represented by a broad range of known and newly discovered RNA species, with many being associated

with 5′ or 3′ regions of genes [4,31,32].

This class includes the well-documented miRNAs, RNAs ~22 nucleotides (nt) long involved in the specific regulation of both

protein-coding, and
putatively non-coding genes,

by post-transcriptional silencing or infrequently
by activation [33-35].

miRNAs serve as major

regulators of gene expression and as
intricate components of the cellular gene expression network [33-38].

Another newly described subclass are the transcription initiation RNAs (tiRNAs), which are

the smallest functional RNAs at only 18 nt in length [39,40].

small ncRNAs classes, including miRNAs, have established roles in tumorigenesis, an intriguing association between
the aberrant expression of ncRNA satellite repeats and cancer has been recently demonstrated [41-46].

Types of human non-coding RNAs

In contrast to miRNAs, lncRNAs, the focus of this article, are

mRNA-like transcripts ranging in length from 200 nt to ~100 kilobases (kb) lacking significant open reading frames.

Many identified lncRNAs are transcribed by RNA polymerase II (RNA pol II) and are polyadenylated, but this is not a fast rule [47,48].
There are examples of lncRNAs, such as the

antisense asOct4-pg5 or the
brain-associated BC200,
- which are functional, but not polyadenylated [49-51].

lncRNA expression levels appear to be lower than protein-coding genes [52-55], and some
lncRNAs are preferentially expressed in specific tissues [21].

Novel lncRNAs may contribute a significant portion of the aforementioned ‘dark matter’ of the human transcriptome [56,57]. In an exciting report
by Kapranov et.al., it was revealed the bulk of the relative mass of RNA in a human cell, exclusive of the ribosomal and mitochondrial RNA,
is represented by non-coding transcripts with no known function [57].

Like miRNAs and protein-coding genes, some

transcriptionally active lncRNA genes display
histone H3K4 trimethylation at their 5′-end and
histone H3K36 trimethylation in the body of the gene [8,58,59].

The small number of characterized human lncRNAs have been associated with a spectrum of biological processes, for example,

epigenetics,
alternative splicing,
nuclear import,

as structural components,
as precursors to small RNAs and
even as regulators of mRNA decay [4,60-70].

Furthermore, accumulating reports of misregulated lncRNA expression across numerous cancer types suggest that

aberrant lncRNA expression may be a major contributor to tumorigenesis [71].

This surge in publications reflects the increasing attention to this subject and a number of useful lncRNA databases have been created .
In this review we highlight the emerging

functional role of aberrant lncRNA expression, including
transcribed ultraconserved regions (T-UCRs), within human carcinomas.

Publications describing cancer-associated ncRNAs. Entries are based on a National Library of Medicine Pubmed search using the terms
“ncRNA” or “non-coding RNA” or “noncoding RNA” or non-protein-coding RNA” with cancer and annual (Jan.1-Dec.31) date limitations. …
Publically available long non-coding RNA online databases

The definition ‘non-coding RNA’ is typically used to describe transcripts where

sequence analysis has failed to identify an open reading frame.

There are cases where ‘non-coding’ transcripts were found to encode short, functional peptides [72]. Currently, a
universal classification scheme to define lncRNAs does not exist. Terms such as

large non-coding RNA,
mRNA-like long RNA, and
intergenic RNA

all define cellular RNAs, exclusive of rRNAs,

greater than 200 nt in length and having no obvious protein-coding capacity [62].

This has led to confusion in the literature as to exactly which transcripts should constitute a lncRNA. One subclass of lncRNAs is called
large or long intergenic ncRNAs (lincRNAs). These lncRNAs are

exclusively intergenic and are
marked by a chromatin signature indicative of transcription [8,58].

RNA species that are bifunctional preclude categorization into either group of

protein-coding or
ncRNAs as

their transcripts function both at the RNA and protein levels [73].

The term ‘lncRNA‘ is used only to describe transcripts with no protein-coding capacity. In the meantime, and for the purposes of this review,
we will consider lncRNAs as a blanket term to encompass

mRNA-like ncRNAs,
lincRNAs, as well as
antisense and intron-encoded transcripts,
T-UCRs and
transcribed pseudogenes.

Discovery of LncRNAs

The earliest reports describing lncRNA predated the discovery of miRNAs, although the term ‘lncRNA‘ had not been coined at the time .
One of the first lncRNA genes reported was the imprinted H19 gene, which was quickly followed by the discovery of the

silencing X-inactive-specific transcript (XIST) lncRNA gene, which
- plays a critical function in X-chromosome inactivation [74,75].

The discovery of the first miRNA lin-14 dramatically redirected the focus of ncRNA research from long ncRNAs to miRNAs [76], and
the discovery of miRNAs revealed RNA could

regulate gene expression and
entire gene networks could be affected by ncRNA expression and

Within the last decade miRNAs were discovered to be associated with cancer. At the time of this writing there are approximately
1049 human miRNAs described in miRBase V16 [80,81] with the potential of

affecting the expression of approximately 60% of protein -coding genes [82,83].

Conversely, the variety and dynamics of lncRNA expression was not to be fully appreciated until the introduction of whole transcriptome sequencing.
With the advent of the FANTOM and ENCODE transcript mapping projects, it was revealed that the mammalian genome is extensively transcribed,
although a large portion of this represented non-coding sequences [3,84]. Coupled with the novel functional annotation of a few lncRNAs, this discovery
promoted research focusing on lncRNA discovery and characterization. Recent reports have described new lncRNA classes such as lincRNAs and T-UCRs [8,58,85].
Current estimates of the lncRNA gene content in the human genome ranges from ~7000 – 23,000 unique lncRNAs, implying this class of ncRNA will
represent an enormous, yet undiscovered, component of normal cellular networks that may be disrupted in cancer biology [62].

Emerging Role of Long Non-Coding RNA in Tumorigenesis

A role for differential lncRNA expression in cancer had been suspected for many years, however, lacked strong supporting evidence [86]. With advancements
in cancer transcriptome profiling and accumulating evidence supporting lncRNA function, a number of differentially expressed lncRNAs have been associated
with cancer. LncRNAs have been implicated to

regulate a range of biological functions and
the disruption of some of these functions, such as
- genomic imprinting and transcriptional regulation,
- plays a critical role in cancer development.

Here we describe some of the better characterized lncRNAs that have been associated with cancer biology.

Human cancer-associated lncRNAs

Imprinted lncRNA genes

Imprinting is a process whereby the copy of a gene inherited from one parent is epigenetically silenced [87,88]. Intriguingly, imprinted regions often
include multiple maternal and paternally expressed genes with a high frequency of ncRNA genes. The imprinted ncRNA genes are implicated in the
imprinting of the region by a variety of mechanisms including

enhancer competition and chromatin remodeling [89].

A key feature of cancer is the loss of this imprinting resulting in altered gene expression [90,91]. Two of the best known imprinted genes
are in fact lncRNAs.

H19

The H19 gene encodes a 2.3 kb lncRNA that is expressed exclusively from the maternal allele. H19 and its reciprocally imprinted protein-coding neighbor
the Insulin-Like Growth Factor 2 or IGF2 gene at 11p15.5 were among the first genes, non-coding or otherwise, found to demonstrate genomic imprinting [74,92].

The expression of H19 is high during vertebrate embryo development, but is

downregulated in most tissues shortly after birth with the exception of skeletal tissue and cartilage [20,93,94].
Loss of imprinting and subsequent strong gene expression has been well-documented in human cancers. Likewise,
loss of imprinting at the H19 locus resulted in high H19 expression in cancers of the esophagus, colon, liver, bladder and with hepatic metastases [95-97].

H19 has been implicated as having both oncogenic and tumor suppression properties in cancer. H19 is upregulated in a number of human cancers, including
hepatocellular, bladder and breast carcinomas, suggesting an oncogenic function for this lncRNA [97-99]. In colon cancer H19 was shown to be directly activated
by the oncogenic transcription factor c-Myc, suggesting

H19 may be an intermediate functionary between c-Myc and downstream gene expression [98].

Conversely, the tumor suppressor gene and transcriptional activator p53 has been shown to

down-regulate H19 expression [100,101].

H19 transcripts also serve as a precursor for miR-675, a miRNA involved in the regulation of developmental genes [102].
miR-675 is processed from the first exon of H19 and functionally

downregulates the tumor suppressor gene retinoblastoma (RB1) in human colorectal cancer, further implying an oncogenic role for H19 [103].

There is evidence suggesting H19 may also play a role in tumor suppression [104,105]. Using a mouse model for colorectal cancer, it was shown that
mice lacking H19 manifested an increased polyp count compared to wild-type [106]. Secondly, a mouse teratocarcinoma model demonstrated larger
tumor growth when the embryo lacked H19, and finally in a hepatocarcinoma model, mice developed cancer much earlier when H19 was absent [107].
The discrepancy as to whether H19 has oncogenic or tumor suppressive potential may be due in part to the bifunctional nature of the lncRNA or may
be context dependent. In either case, the precise functional and biological role of H19 remains to be determined.

XIST – X-inactive-specific transcript

The 17 kb lncRNA XIST is arguably an archetype for the study of functional lncRNAs in mammalian cells, having been studied for nearly two decades.
In female cells, the XIST transcript plays a critical role in X-chromosome inactivation by

physically coating one of the two X-chromosomes, and is necessary for the
cis-inactivation of the over one thousand X-linked genes [75,108-110].

Like the lncRNAs HOTAIR and ANRIL, XIST associates with polycomb-repressor proteins, suggesting

a common pathway of inducing silencing utilized by diverse lncRNAs.

Discovery of Molecular Mechanisms of Traditional Chinese Medicinal Formula Si-Wu-Tang Using Gene Expression Microarray and Connectivity Map
by Zhining Wen, Zhijun Wang, Steven Wang, Ranadheer Ravula, Lun Yang, …et al.
PLoS ONE (2011); 6:(3), Publisher: PLoS, Pages: 14 http.//dx.doi.org/10.1371/journal.pone.0018278 PubMed: 21464939
http://dx.plos.org/Wen Z, Wang Z, Wang Z, et al./discovery of molecular mechanisms of traditional chinese medicinal formula…/
To pursue a systematic approach to discovery of mechanisms of action of traditional Chinese medicine (TCM), we used

microarrays,
bioinformatics and the
Connectivity Map (CMAP)
- to examine TCM-induced changes in gene expression.

We demonstrated that this approach can be used to elucidate new molecular targets using a model TCM herbal formula Si-Wu-Tang (SWT) which is

widely used for women’s health.

The human breast cancer MCF-7 cells treated with 0.1 µM estradiol or 2.56 mg/ml of SWT

showed dramatic gene expression changes, while
no significant change was detected for ferulic acid, a known bioactive compound of SWT.

Pathway analysis using

differentially expressed genes related to the treatment effect
identified that expression of genes in the nuclear factor erythroid 2-related factor 2 (Nrf2) cytoprotective pathway
was most significantly affected by SWT,
- but not by estradiol or ferulic acid.
The Nrf2-regulated genes
- HMOX1,
- GCLC,
- GCLM,
- SLC7A11 and
- NQO1 were
upreguated by SWT in a dose-dependent manner, which was validated by real-time RT-PCR. Consistently,
treatment with SWT and its four herbal ingredients resulted in an
increased antioxidant response element (ARE)-luciferase reporter activity in MCF-7 and HEK293 cells.