Sinhcaf-dependent histone deacetylation is essential for primordial germ cell specification
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ABSTRACT:
SUBMITTER: Dr. Binbin Tao
PROVIDER: S-SCDT-EMBOR-2021-54387-T | biostudies-other |
REPOSITORIES: biostudies-other
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Project description:Germline specification is a fundamental step for human reproduction and this biological phenomenon possesses technical challenges to study in vivo as it occurs immediately after blastocyst implantation. The establishment of in vitro human primordial germ cell-like cells (hPGCLCs) induction system allows sophisticated characterization of human primordial germ cells (hPGCs) development. However, the underlying molecular mechanisms of hPGCLC specification are not fully elucidated. Here, we observed particularly high expression of the histone demethylase KDM2B in male fetal germ cells (FGCs) but not in male somatic cells. Besides, KDM2B shared similar expression pattern with hPGC marker genes in hPGCLCs, suggesting an important role of KDM2B in germ cell development. Although deletion of KDM2B had no significant effects on human embryonic stem cell (hESC)'s pluripotency, loss of KDM2B dramatically impaired hPGCLCs differentiation whereas ectopically expressed KDM2B could efficiently rescue such defect, indicating this defect was due to KDM2B's loss in hPGCLC specification. Mechanistically, as revealed by the transcriptional profiling, KDM2B suppressed the expression of somatic genes thus inhibited somatic differentiation during hPGCLC specification. These data collectively indicate that KDM2B is an indispensable epigenetic regulator for hPGCLC specification, shedding lights on how epigenetic regulations orchestrate transcriptional events in hPGC development for future investigation.
| S-EPMC7893587 | biostudies-literature
Project description:BackgroundPrimordial germ cell (PGC) specification is the first crucial step in germ line development. However, owing to significant challenges regarding the in vivo system, such as the complex cellular environment and potential problems with embryo manipulation, it is desirable to generate embryonic stem (ES) cells that are capable of overcoming these aforementioned limitations in order to provide a potential in vitro model to recapitulate the developmental processes in vivo.Methodology and principal findingsHere, we studied the detailed process of PGC specification from stella-GFP ES cells. We first observed the heterogeneous expression of stella in ES cells. However, neither Stella-positive ES cells nor Stella-negative ES cells shared a similar gene expression pattern with either PGCs or PGC precursors. Second, we derived PGCs from ES cells using two differentiation methods, namely the attachment culture technique and the embryoid body (EB) method. Compared with PGCs derived via the attachment culture technique, PGCs derived via the EB method that had undergone the sequential erasure of Peg3 followed by Igf2r resulted in a cell line in which the expression dynamics of T, Fgf8 and Sox17, in addition to the expression of the epiblast markers, were more similar to the in vivo expression, thus demonstrating that the process of PGC derivation was more faithfully recapitulated using the EB method. Furthermore, we developed an in vitro model of PGC specification in a completely chemically defined medium (CDM) that indicated that BMP4 and Wnt3a promoted PGC derivation, whereas BMP8b and activinA had no observable effect on PGC derivation.Conclusions and significanceThe in vitro model we have established can recapitulate the developmental processes in vivo and provides new insights into the mechanism of PGC specification.
| S-EPMC2602984 | biostudies-literature
Project description:Our current knowledge of the mechanisms leading to human primordial germ cell (PGC) specification stems solely from differentiation experiments starting from human pluripotent stem cells. However, information regarding the origin of PGCs in vivo remains obscure. Here we apply an improved system for extended in vitro culture of human embryos to investigate the presence of PGC-like cells (PGCLCs) 12 days post fertilization (dpf). Good quality blastocysts (n?=?141) were plated at 6 dpf and maintained in hypoxia, in medium supplemented with Activin A until 12 dpf. We primarily reveal that 12 dpf outgrowths recapitulate human peri-implantation events and demonstrate that blastocyst quality significantly impacts both embryo viability at 12 dpf, as well as the presence of POU5F1+ cells within viable outgrowths. Moreover, detailed examination of 12 dpf blastocyst outgrowths revealed a population of POU5F1+, SOX2- and SOX17+ cells that may correspond to PGCLCs, alongside POU5F1+ epiblast-like cells and GATA6+ endoderm-like cells. Our findings suggest that, in human, PGC precursors may become specified within the epiblast and migrate either transiently to the extra-embryonic mesoderm or directly to the dorsal part of the yolk sac endoderm around 12 dpf. This is a descriptive analysis and as such the conclusion that POU5F1+ and SOX17+ cells represent bona fide PGCs can only be considered as preliminary. In the future, other PGC markers may be used to further validate the observed cell populations. Overall, our findings provide insights into the origin of the human germline and may serve as a foundation to further unravel the molecular mechanisms governing PGC specification in human.
| S-EPMC6802404 | biostudies-literature
Project description:Specification of the germ cell lineage is required for sexual reproduction in animals. The mechanism of germ cell specification varies among animals but roughly clusters into either inherited or inductive mechanisms. The inductive mechanism, the use of cell-cell interactions for germ cell specification, appears to be the ancestral mechanism in animal phylogeny, yet the pathways responsible for this process are only recently surfacing. Here, we show that germ cell factors in the sea star initially are present broadly, then become restricted dorsally and then in the left side of the embryo where the germ cells form a posterior enterocoel. We find that Nodal signaling is required for the restriction of two germ cell factors, Nanos and Vasa, during the early development of this animal. We learned that Nodal inhibits germ cell factor accumulation in three ways including: inhibition of specific transcription, degradation of specific mRNAs and inhibition of tissue morphogenesis. These results document a signaling mechanism required for the sequential restriction of germ cell factors, which causes a specific set of embryonic cells to become the primordial germ cells.
| S-EPMC5825842 | biostudies-other
Project description:The localization and organization of mitochondria- and ribonucleoprotein granule-rich germ plasm is essential for many aspects of germ cell development. In Xenopus, germ plasm is maternally inherited and is required for the specification of primordial germ cells (PGCs). Germ plasm is aggregated into larger patches during egg activation and cleavage and is ultimately translocated perinuclearly during gastrulation. Although microtubule dynamics and a kinesin (Kif4a) have been implicated in Xenopus germ plasm localization, little is known about how germ plasm distribution is regulated. Here, we identify a role for maternal Xenopus Syntabulin in the aggregation of germ plasm following fertilization. We show that depletion of sybu mRNA using antisense oligonucleotides injected into oocytes results in defects in the aggregation and perinuclear transport of germ plasm and subsequently in reduced PGC numbers. Using live imaging analysis, we also characterize a novel role for Sybu in the collection of germ plasm in vegetal cleavage furrows by surface contraction waves. Additionally, we show that a localized kinesin-like protein, Kif3b, is also required for germ plasm aggregation and that Sybu functionally interacts with Kif3b and Kif4a in germ plasm aggregation. Overall, these data suggest multiple coordinate roles for kinesins and adaptor proteins in controlling the localization and distribution of a cytoplasmic determinant in early development.
| S-EPMC5714693 | biostudies-literature
Project description:During early mammalian development, the pluripotent cells of the embryo are exposed to a combination of signals that drive exit from pluripotency and germ layer differentiation. At the same time, a small population of pluripotent cells give rise to the primordial germ cells (PGCs), the precursors of the sperm and egg, which pass on heritable genetic information to the next generation. Despite the importance of PGCs, it remains unclear how they are first segregated from the soma, and if this involves distinct responses to their signaling environment. To investigate this question, we mapped BMP, MAPK and WNT signaling responses over time in PGCs and their surrounding niche in vitro and in vivo at single-cell resolution. We showed that, in the mouse embryo, early PGCs exhibit lower BMP and MAPK responses compared to neighboring extraembryonic mesoderm cells, suggesting the emergence of distinct signaling regulatory mechanisms in the germline versus soma. In contrast, PGCs and somatic cells responded comparably to WNT, indicating that this signal alone is not sufficient to promote somatic differentiation. Finally, we investigated the requirement of a BMP response for these cell fate decisions. We found that cell lines with a mutation in the BMP receptor (Bmpr1a-/-), which exhibit an impaired BMP signaling response, can efficiently generate PGC-like cells revealing that canonical BMP signaling is not cell autonomously required to direct PGC-like differentiation.
| S-EPMC8186728 | biostudies-literature
Project description:Transitions in cell states are controlled by combinatorial actions of transcription factors. BLIMP1, the key regulator of primordial germ cell (PGC) specification, apparently acts together with PRDM14 and AP2?. To investigate their individual and combinatorial functions, we first sought an in vitro system for transcriptional readouts and chromatin immunoprecipitation sequencing analysis. We then integrated this data with information from single-cell transcriptome analysis of normal and mutant PGCs. Here we show that BLIMP1 binds directly to repress somatic and cell proliferation genes. It also directly induces AP2?, which together with PRDM14 initiates the PGC-specific fate. We determined the occupancy of critical genes by AP2?-which, when computed altogether with those of BLIMP1 and PRDM14 (both individually and cooperatively), reveals a tripartite mutually interdependent transcriptional network for PGCs. We also demonstrate that, in principle, BLIMP1, AP2? and PRDM14 are sufficient for PGC specification, and the unprecedented resetting of the epigenome towards a basal state.
| S-EPMC3796875 | biostudies-literature
Project description:In animals, primordial germ cells (PGCs) give rise to the germ lines, the cell lineages that produce sperm and eggs. PGCs form in embryogenesis, typically by one of two modes: a likely ancestral mode wherein germ cells are induced during embryogenesis by cell-cell signaling (induction) or a derived mechanism whereby germ cells are specified by using germ plasm-that is, maternally specified germ-line determinants (inheritance). The causes of the shift to germ plasm for PGC specification in some animal clades remain largely unknown, but its repeated convergent evolution raises the question of whether it may result from or confer an innate selective advantage. It has been hypothesized that the acquisition of germ plasm confers enhanced evolvability, resulting from the release of selective constraint on somatic gene networks in embryogenesis, thus leading to acceleration of an organism's protein-sequence evolution, particularly for genes expressed at early developmental stages, and resulting in high speciation rates in germ plasm-containing lineages (denoted herein as the "PGC-specification hypothesis"). Although that hypothesis, if supported, could have major implications for animal evolution, our recent large-scale coding-sequence analyses from vertebrates and invertebrates provided important examples of genera that do not support the hypothesis of liberated constraint under germ plasm. Here, we consider reasons why germ plasm might be neither a direct target of selection nor causally linked to accelerated animal evolution. We explore alternate scenarios that could explain the repeated evolution of germ plasm and propose potential consequences of the inheritance and induction modes to animal evolutionary biology.
| S-EPMC5468662 | biostudies-literature
Project description:BackgroundPrimordial germ cells (PGCs) give rise to gametes that are responsible for the development of a new organism in the next generation. Two modes of germ line specification have been described: the inheritance of asymmetrically-localized maternally provided cytoplasmic determinants and the induction of the PGC fate by other cell types. PGCs specification in zebrafish appears to depend on inheritance of germ plasm in which several RNA molecules such as vasa and nanos reside. Whether the specification mode of PGCs found in zebrafish is general for other fish species was brought into question upon analysis of olvas expression--the vasa homologue in another teleost, medaka (Oryzias latipes). Here, in contrast to the findings in zebrafish, the PGCs are found in a predictable position relative to a somatic structure, the embryonic shield. This finding, coupled with the fact that vasa mRNA, which is localized to the germ plasm of zebrafish but does not label a similar structure in medaka opened the possibility of fundamentally different mechanisms governing PGC specification in these two fish species.ResultsIn this study we addressed the question concerning the mode of PGC specification in medaka using embryological experiments, analysis of RNA stability in the PGCs and electron microscopy observations. Dramatic alterations in the somatic environment, i.e. induction of a secondary axis or mesoderm formation alteration, did not affect the PGC number. Furthermore, the PGCs of medaka are capable of protecting specific RNA molecules from degradation and could therefore exhibit a specific mRNA expression pattern controlled by posttrancriptional mechanisms. Subsequent analysis of 4-cell stage medaka embryos using electron microscopy revealed germ plasm-like structures located at a region corresponding to that of zebrafish germ plasm.ConclusionTaken together, these results are consistent with the idea that in medaka the inheritance of maternally provided asymmetrically-localized cytoplasmic determinants directs cells to assume the germ line fate similar to zebrafish PGCs.
| S-EPMC1781431 | biostudies-literature
Project description:Primordial germ cells (PGCs) are common ancestors of all germline cells. However, mechanistic understanding of how PGC specification occurs is limited. Here, we identified transcription factor CP2-like 1 (Tfcp2l1), an important pluripotency factor, as a pivotal factor for PGC-like cell (PGCLC) specification. High-throughput sequencing and quantitative real-time PCR analysis showed that Tfcp2l1 expression is gradually increased during mouse and human epiblast differentiation into PGCLCs in vivo and in vitro. Consequently, overexpression of Tfcp2l1 can enhance the specification efficiency even without inductive cytokines in mouse epiblast-like cells derived from embryonic stem cells, while knockdown of Tfcp2l1 significantly inhibits PGCLC generation. Mechanistic studies revealed that Tfcp2l1 exerts its function partially through the direct induction of PR domain zinc finger protein 14, a key PGC marker, as downregulation of the PR domain zinc finger protein 14 transcript can impair the ability of Tfcp2l1 to direct PGCLC commitment. Importantly, we finally demonstrated that the crucial role of the human homolog Tfcp2l1 in promoting PGCLC specification is conserved in human pluripotent stem cells. Together, our data uncover a novel function of Tfcp2l1 in PGCLC fate determination and facilitate a better understanding of germ cell development.
| S-EPMC8517209 | biostudies-literature