Reporter and Curator: Dr. Sudipta Saha, Ph.D.
Gametogenesis is a biological process by which precursor cells undergo cell division and differentiation to form mature haploid gametes. Human gametogenesis occurs by mitotic division of gametogonia, followed by meiotic division of gametocytes into various gametes. During this process, the gamete genome experiences both programmed and spontaneous changes, among which meiotic recombination shuffles the two haploid somatic genomes to create a unique hybrid haploid genome for each gamete cell, while accumulated replication errors contribute point mutations that may affect the gametes’ functionality. This results in an enormous variety of new genomes being created in the gametes, thereby enabling one’s children to add to the genetic diversity of the human race in a more complex manner than by simply mixing and matching entire parental chromosomes. The genome-wide recombination activity and de novo mutation rate have been directly characterized in many model organisms. However, it has been unclear how an individual human’s genome is edited during gametogenesis. Despite the advances in personal genomics, gamete genome variation within individuals, especially fine-scale personal recombination activity and germline mutation rates, has been as yet generally inaccessible.
An important feature of single molecule multiple displacement amplification (MDA) is its repetitive usage of the originating genuine template molecule. Even if an amplification error happens in the initial stage, there will still be a large fraction of products preserving the correct base information from the original template, and the power of statistics from multiple coverage discriminates these errors from true genomic variation. Using this microfluidic MDA approach, for the first genome-wide single-cell analysis of human sperm was reported. A personal recombination map was created for an individual to measure the rate of de novo mutations in this individual’s germline. The advantage of sampling a large set of meioses from a single individual for fine-scale analysis allowed to uncover individual specific features potentially buried under population data. It was proposed that this partially overlapping feature is also the general pattern in individuals. While some hot spots are dying in some people, new recombination activities evolve to refill the hot spot pool. Support for this theory comes from single-cell analysis. Recombination data from 91 single sperm cells presented a comprehensive landscape of personal recombination activity. Genome-wide meiotic drive and gene conversion were also directly tested. Single-cell whole-genome sequencing further revealed primary information about human sperm genome instability and mutation rate. In this study, microfluidics to single-cell whole genome amplification was applied. This technique not only enabled great parallelization, but also improved amplification performance. MDA is sensitive to environmental contamination, and extensive sample purification is required for traditional bench-top whole genome amplifications.
The data from this study suggested that the germline mutation rate can vary greatly among different individuals, but not among different cells from the same individual. This may explain why the male mutation rate is not always higher than the female. DNA methylation also affects genome instability and C/T point mutation levels but in opposite ways. A fine tuned methylation level is therefore required for high-quality sperm genome. The ability to study a large number of single sperm cells has offered several new insights in meiosis. Studying the germline genome is but one application of single-cell genomics, and it is expected that the method will find applications in many other fields, including cancer, aging, immunology, and developmental biology.
Source References:
Genome-wide Single-Cell Analysis of Recombination Activity and De Novo Mutation Rates in Human Sperm.
Personal Recombination Map from Individual’s Sperm Cell and its Importance
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Dr. Saha,
Thank you for another great post.
Transferability of knowledge from the germline genome as an example of single-cell genomics, is most important for potential finding of new applications in many other fields, including cancer, aging, immunology, and developmental biology.
Harvesting that type of cell is easier that harvesting a bone marrow cell, though due to the chromosome structure of reproductive cell one may envision limitations on the transferability of knowledge. What do you say?
PUT IT IN CONTEXT OF CANCER CELL MOVEMENT
The contraction of skeletal muscle is triggered by nerve impulses, which stimulate the release of Ca2+ from the sarcoplasmic reticuluma specialized network of internal membranes, similar to the endoplasmic reticulum, that stores high concentrations of Ca2+ ions. The release of Ca2+ from the sarcoplasmic reticulum increases the concentration of Ca2+ in the cytosol from approximately 10-7 to 10-5 M. The increased Ca2+ concentration signals muscle contraction via the action of two accessory proteins bound to the actin filaments: tropomyosin and troponin (Figure 11.25). Tropomyosin is a fibrous protein that binds lengthwise along the groove of actin filaments. In striated muscle, each tropomyosin molecule is bound to troponin, which is a complex of three polypeptides: troponin C (Ca2+-binding), troponin I (inhibitory), and troponin T (tropomyosin-binding). When the concentration of Ca2+ is low, the complex of the troponins with tropomyosin blocks the interaction of actin and myosin, so the muscle does not contract. At high concentrations, Ca2+ binding to troponin C shifts the position of the complex, relieving this inhibition and allowing contraction to proceed.
Figure 11.25
Association of tropomyosin and troponins with actin filaments. (A) Tropomyosin binds lengthwise along actin filaments and, in striated muscle, is associated with a complex of three troponins: troponin I (TnI), troponin C (TnC), and troponin T (TnT). In (more ) Contractile Assemblies of Actin and Myosin in Nonmuscle Cells
Contractile assemblies of actin and myosin, resembling small-scale versions of muscle fibers, are present also in nonmuscle cells. As in muscle, the actin filaments in these contractile assemblies are interdigitated with bipolar filaments of myosin II, consisting of 15 to 20 myosin II molecules, which produce contraction by sliding the actin filaments relative to one another (Figure 11.26). The actin filaments in contractile bundles in nonmuscle cells are also associated with tropomyosin, which facilitates their interaction with myosin II, probably by competing with filamin for binding sites on actin.
Figure 11.26
Contractile assemblies in nonmuscle cells. Bipolar filaments of myosin II produce contraction by sliding actin filaments in opposite directions. Two examples of contractile assemblies in nonmuscle cells, stress fibers and adhesion belts, were discussed earlier with respect to attachment of the actin cytoskeleton to regions of cell-substrate and cell-cell contacts (see Figures 11.13 and 11.14). The contraction of stress fibers produces tension across the cell, allowing the cell to pull on a substrate (e.g., the extracellular matrix) to which it is anchored. The contraction of adhesion belts alters the shape of epithelial cell sheets: a process that is particularly important during embryonic development, when sheets of epithelial cells fold into structures such as tubes.
The most dramatic example of actin-myosin contraction in nonmuscle cells, however, is provided by cytokinesisthe division of a cell into two following mitosis (Figure 11.27). Toward the end of mitosis in animal cells, a contractile ring consisting of actin filaments and myosin II assembles just underneath the plasma membrane. Its contraction pulls the plasma membrane progressively inward, constricting the center of the cell and pinching it in two. Interestingly, the thickness of the contractile ring remains constant as it contracts, implying that actin filaments disassemble as contraction proceeds. The ring then disperses completely following cell division.
Figure 11.27
Cytokinesis. Following completion of mitosis (nuclear division), a contractile ring consisting of actin filaments and myosin II divides the cell in two.
http://www.ncbi.nlm.nih.gov/books/NBK9961/
This is good. I don’t recall seeing it in the original comment. I am very aware of the actin myosin troponin connection in heart and in skeletal muscle, and I did know about the nonmuscle work. I won’t deal with it now, and I have been working with Aviral now online for 2 hours.
I have had a considerable background from way back in atomic orbital theory, physical chemistry, organic chemistry, and the equilibrium necessary for cations and anions. Despite the calcium role in contraction, I would not discount hypomagnesemia in having a disease role because of the intracellular-extracellular connection. The description you pasted reminds me also of a lecture given a few years ago by the Nobel Laureate that year on the mechanism of cell division.