RESUMO
The specialized cell cycle of meiosis transforms diploid germ cells into haploid gametes. In mammals, diploid spermatogenic cells acquire the competence to initiate meiosis in response to retinoic acid. Previous mouse studies revealed that MEIOC interacts with RNA-binding proteins YTHDC2 and RBM46 to repress mitotic genes and to promote robust meiotic gene expression in spermatogenic cells that have initiated meiosis. Here, we have used the enhanced resolution of scRNA-seq and bulk RNA-seq of developmentally synchronized spermatogenesis to define how MEIOC molecularly supports early meiosis in spermatogenic cells. We demonstrate that MEIOC mediates transcriptomic changes before meiotic initiation, earlier than previously appreciated. MEIOC, acting with YTHDC2 and RBM46, destabilizes its mRNA targets, including the transcriptional repressors E2f6 and Mga, in mitotic spermatogonia. MEIOC thereby derepresses E2F6- and MGA-repressed genes, including Meiosin and other meiosis-associated genes. This confers on spermatogenic cells the molecular competence to, in response to retinoic acid, fully activate the transcriptional regulator STRA8-MEIOSIN, which is required for the meiotic G1/S phase transition and for meiotic gene expression. We conclude that, in mice, mRNA decay mediated by MEIOC-YTHDC2-RBM46 enhances the competence of spermatogenic cells to initiate meiosis.
Assuntos
Meiose , RNA Mensageiro , Proteínas de Ligação a RNA , Espermatogênese , Animais , Masculino , Camundongos , RNA Mensageiro/metabolismo , RNA Mensageiro/genética , Espermatogênese/genética , Espermatogênese/fisiologia , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Espermatogônias/metabolismo , Espermatogônias/citologia , Tretinoína/metabolismo , Tretinoína/farmacologia , Estabilidade de RNA/genética , Regulação da Expressão Gênica no Desenvolvimento , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , RNA HelicasesRESUMO
Despite numerous female contraceptive options, nearly half of all pregnancies are unintended. Family planning choices for men are currently limited to unreliable condoms and invasive vasectomies with questionable reversibility. Here, we report the development of an oral contraceptive approach based on transcriptional disruption of cyclical gene expression patterns during spermatogenesis. Spermatogenesis involves a continuous series of self-renewal and differentiation programs of spermatogonial stem cells (SSCs) that is regulated by retinoic acid (RA)-dependent activation of receptors (RARs), which control target gene expression through association with corepressor proteins. We have found that the interaction between RAR and the corepressor silencing mediator of retinoid and thyroid hormone receptors (SMRT) is essential for spermatogenesis. In a genetically engineered mouse model that negates SMRT-RAR binding (SMRTmRID mice), the synchronized, cyclic expression of RAR-dependent genes along the seminiferous tubules is disrupted. Notably, the presence of an RA-resistant SSC population that survives RAR de-repression suggests that the infertility attributed to the loss of SMRT-mediated repression is reversible. Supporting this notion, we show that inhibiting the action of the SMRT complex with chronic, low-dose oral administration of a histone deacetylase inhibitor reversibly blocks spermatogenesis and fertility without affecting libido. This demonstration validates pharmacologic targeting of the SMRT repressor complex for non-hormonal male contraception.
Assuntos
Proteínas de Ligação a DNA , Proteínas Repressoras , Humanos , Feminino , Masculino , Animais , Camundongos , Proteínas de Ligação a DNA/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Proteínas Correpressoras/genética , Correpressor 2 de Receptor Nuclear/genética , Tretinoína/farmacologia , Anticoncepção , Correpressor 1 de Receptor NuclearRESUMO
Actin-related proteins (Arps) are classified according to their similarity to actin and are involved in diverse cellular processes. ACTL7B is a testis-specific Arp, and is highly conserved in rodents and primates. ACTL7B is specifically expressed in round and elongating spermatids during spermiogenesis. Here, we have generated an Actl7b-null allele in mice to unravel the role of ACTL7B in sperm formation. Male mice homozygous for the Actl7b-null allele (Actl7b-/-) were infertile, whereas heterozygous males (Actl7b+/-) were fertile. Severe spermatid defects, such as detached acrosomes, disrupted membranes and flagella malformations start to appear after spermiogenesis step 9 in Actl7b-/- mice, finally resulting in spermatogenic arrest. Abnormal spermatids were degraded and levels of autophagy markers were increased. Co-immunoprecipitation with mass spectrometry experiments identified an interaction between ACTL7B and the LC8 dynein light chains DYNLL1 and DYNLL2, which are first detected in step 9 spermatids and mislocalized when ACTL7B is absent. Our data unequivocally establish that mutations in ACTL7B are directly related to male infertility, pressing for additional research in humans.
Assuntos
Actinas , Dineínas , Animais , Humanos , Masculino , Camundongos , Actinas/metabolismo , Dineínas do Citoplasma/metabolismo , Dineínas/genética , Dineínas/metabolismo , Sêmen/metabolismo , Espermátides/metabolismo , Espermatogênese/genética , Espermatozoides/metabolismo , Testículo/metabolismoRESUMO
The special cell cycle known as meiosis transforms diploid germ cells into haploid gametes. In mammalian testes, diploid spermatogenic cells become competent to transition from mitosis to meiosis in response to retinoic acid. In mice, previous studies revealed that MEIOC, alongside binding partners YTHDC2 and RBM46, represses mitotic genes and promotes robust meiotic gene expression in spermatogenic cells that have already initiated meiosis. Here, we molecularly dissect MEIOC-dependent regulation in mouse spermatogenic cells and find that MEIOC actually shapes the transcriptome much earlier, even before meiotic initiation. MEIOC, acting with YTHDC2 and RBM46, destabilizes mRNA targets, including transcriptional repressors E2f6 and Mga, in mitotic spermatogonia. MEIOC thereby derepresses E2F6- and MGA-repressed genes, including Meiosin and other meiosis-associated genes. This confers on spermatogenic cells the molecular competence to, in response to retinoic acid, fully activate the STRA8-MEIOSIN transcriptional regulator, which is required for the meiotic G1/S cell cycle transition and meiotic gene expression. We conclude that in mice, mRNA decay mediated by MEIOC-YTHDC2-RBM46 enhances the competence of mitotic spermatogonia to transit from mitosis to meiosis.
RESUMO
Polycomb group (PcG) proteins maintain the repressed state of lineage-inappropriate genes and are therefore essential for embryonic development and adult tissue homeostasis. One critical function of PcG complexes is modulating chromatin structure. Canonical Polycomb repressive complex 1 (cPRC1), particularly its component CBX2, can compact chromatin and phase-separate in vitro. These activities are hypothesized to be critical for forming a repressed physical environment in cells. While much has been learned by studying these PcG activities in cell culture models, it is largely unexplored how cPRC1 regulates adult stem cells and their subsequent differentiation in living animals. Here, we show in vivo evidence of a critical nonenzymatic repressive function of cPRC1 component CBX2 in the male germline. CBX2 is up-regulated as spermatogonial stem cells differentiate and is required to repress genes that were active in stem cells. CBX2 forms condensates (similar to previously described Polycomb bodies) that colocalize with target genes bound by CBX2 in differentiating spermatogonia. Single-cell analyses of mosaic Cbx2 mutant testes show that CBX2 is specifically required to produce differentiating A1 spermatogonia. Furthermore, the region of CBX2 responsible for compaction and phase separation is needed for the long-term maintenance of male germ cells in the animal. These results emphasize that the regulation of chromatin structure by CBX2 at a specific stage of spermatogenesis is critical, which distinguishes this from a mechanism that is reliant on histone modification.
Assuntos
Núcleo Celular , Cromatina , Animais , Masculino , Cromatina/metabolismo , Núcleo Celular/metabolismo , Complexo Repressor Polycomb 1/genética , Complexo Repressor Polycomb 1/metabolismo , Proteínas do Grupo Polycomb/genética , Proteínas do Grupo Polycomb/metabolismo , Espermatogênese/genéticaRESUMO
In brief: The testis-specific transcription factor, TCFL5, expressed in pachytene spermatocytes regulates the meiotic gene expression program in collaboration with the transcription factor A-MYB. Abstract: In male mice, the transcription factors STRA8 and MEISON initiate meiosis I. We report that STRA8/MEISON activates the transcription factors A-MYB and TCFL5, which together reprogram gene expression after spermatogonia enter into meiosis. TCFL5 promotes the transcription of genes required for meiosis, mRNA turnover, miR-34/449 production, meiotic exit, and spermiogenesis. This transcriptional architecture is conserved in rhesus macaque, suggesting TCFL5 plays a central role in meiosis and spermiogenesis in placental mammals. Tcfl5em1/em1 mutants are sterile, and spermatogenesis arrests at the mid- or late-pachytene stage of meiosis. Moreover, Tcfl5+/em1 mutants produce fewer motile sperm.
Assuntos
Placenta , Fatores de Transcrição , Animais , Feminino , Masculino , Camundongos , Gravidez , Macaca mulatta/metabolismo , Mamíferos/metabolismo , Meiose , Placenta/metabolismo , Sêmen/metabolismo , Espermatócitos/metabolismo , Espermatogênese/genética , Testículo/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Primary Ovarian Insufficiency (POI) is a major cause of infertility, but its etiology remains poorly understood. Using whole-exome sequencing in a family with three cases of POI, we identified the candidate missense variant S167L in HSF2BP, an essential meiotic gene. Functional analysis of the HSF2BP-S167L variant in mouse showed that it behaves as a hypomorphic allele compared to a new loss-of-function (knock-out) mouse model. Hsf2bpS167L/S167L females show reduced fertility with smaller litter sizes. To obtain mechanistic insights, we identified C19ORF57/BRME1 as a strong interactor and stabilizer of HSF2BP and showed that the BRME1/HSF2BP protein complex co-immunoprecipitates with BRCA2, RAD51, RPA and PALB2. Meiocytes bearing the HSF2BP-S167L variant showed a strongly decreased staining of both HSF2BP and BRME1 at the recombination nodules and a reduced number of the foci formed by the recombinases RAD51/DMC1, thus leading to a lower frequency of crossovers. Our results provide insights into the molecular mechanism of HSF2BP-S167L in human ovarian insufficiency and sub(in)fertility.
Assuntos
Proteínas de Transporte , Proteínas de Choque Térmico , Meiose/genética , Mutação de Sentido Incorreto/genética , Insuficiência Ovariana Primária/genética , Recombinação Genética/genética , Animais , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Feminino , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Camundongos , Camundongos Knockout , Rad51 Recombinase/genética , Rad51 Recombinase/metabolismo , Sequenciamento do ExomaRESUMO
Fertility across metazoa requires the germline-specific DAZ family of RNA-binding proteins. Here we examine whether DAZL directly regulates progenitor spermatogonia using a conditional genetic mouse model and in vivo biochemical approaches combined with chemical synchronization of spermatogenesis. We find that the absence of Dazl impairs both expansion and differentiation of the spermatogonial progenitor population. In undifferentiated spermatogonia, DAZL binds the 3' UTRs of ~2,500 protein-coding genes. Some targets are known regulators of spermatogonial proliferation and differentiation while others are broadly expressed, dosage-sensitive factors that control transcription and RNA metabolism. DAZL binds 3' UTR sites conserved across vertebrates at a UGUU(U/A) motif. By assessing ribosome occupancy in undifferentiated spermatogonia, we find that DAZL increases translation of its targets. In total, DAZL orchestrates a broad translational program that amplifies protein levels of key spermatogonial and gene regulatory factors to promote the expansion and differentiation of progenitor spermatogonia.
Assuntos
Diferenciação Celular , Proteínas de Ligação a RNA , Espermatogênese , Regiões 3' não Traduzidas , Animais , Diferenciação Celular/fisiologia , Masculino , Camundongos , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Espermatogênese/fisiologia , Espermatogônias/metabolismoRESUMO
The journal and the authors apologise for an error in the above titled article published in this journal (vol 144, pp 433445). The authors inadvertently presented duplicate sperm images for XY and XESxrbO mouse testes of Fig. 6 (bottom panels). This error does not change the findings of the paper, as this figure does not give a quantitative breakdown of the proportions of different shapes.
RESUMO
Germ cells undergo many developmental transitions before ultimately becoming either eggs or sperm, and during embryonic development these transitions include epigenetic reprogramming, quiescence, and meiosis. To begin understanding the transcriptional regulation underlying these complex processes, we examined the spatial and temporal expression of TAF4b, a variant TFIID subunit required for fertility, during embryonic germ cell development. By analyzing published datasets and using our own experimental system to validate these expression studies, we determined that both Taf4b mRNA and protein are highly germ cell-enriched and that Taf4b mRNA levels dramatically increase from embryonic day 12.5-18.5. Surprisingly, additional mRNAs encoding other TFIID subunits are coordinately upregulated through this time course, including Taf7l and Taf9b. The expression of several of these germ cell-enriched TFIID genes is dependent upon Dazl and/or Stra8, known regulators of germ cell development and meiosis. Together, these data suggest that germ cells employ a highly specialized and dynamic form of TFIID to drive the transcriptional programs that underlie mammalian germ cell development.
Assuntos
Gametogênese , Regulação da Expressão Gênica no Desenvolvimento , Células Germinativas/metabolismo , Fatores Associados à Proteína de Ligação a TATA/genética , Fator de Transcrição TFIID/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Proteína 1 Suprimida em Azoospermia/genética , Proteína 1 Suprimida em Azoospermia/metabolismo , Células Germinativas/citologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fatores Associados à Proteína de Ligação a TATA/metabolismo , Fator de Transcrição TFIID/metabolismoRESUMO
GCNA proteins are expressed across eukarya in pluripotent cells and have conserved functions in fertility. GCNA homologs Spartan (DVC-1) and Wss1 resolve DNA-protein crosslinks (DPCs), including Topoisomerase-DNA adducts, during DNA replication. Here, we show that GCNA mutants in mouse and C. elegans display defects in genome maintenance including DNA damage, aberrant chromosome condensation, and crossover defects in mouse spermatocytes and spontaneous genomic rearrangements in C. elegans. We show that GCNA and topoisomerase II (TOP2) physically interact in both mice and worms and colocalize on condensed chromosomes during mitosis in C. elegans embryos. Moreover, C. elegans gcna-1 mutants are hypersensitive to TOP2 poison. Together, our findings support a model in which GCNA provides genome maintenance functions in the germline and may do so, in part, by promoting the resolution of TOP2 DPCs.
Assuntos
Replicação do DNA , DNA Topoisomerases Tipo II/metabolismo , Proteínas de Ligação a DNA/metabolismo , Instabilidade Genômica , Mitose , Proteínas Nucleares/metabolismo , Espermatócitos/citologia , Animais , Caenorhabditis elegans , Dano ao DNA , Reparo do DNA , DNA Topoisomerases Tipo II/genética , Proteínas de Ligação a DNA/genética , Genoma , Células Germinativas , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mutação , Proteínas Nucleares/genética , Espermatócitos/metabolismo , EspermatogêneseRESUMO
Retinoic acid (RA), a derivative of vitamin A, is critical for the production of oocytes and sperm in mammals. These gametes derive from primordial germ cells, which colonize the nascent gonad, and later undertake sexual differentiation to produce oocytes or sperm. During fetal development, germ cells in the ovary initiate meiosis in response to RA, whereas those in the testis do not yet initiate meiosis, as they are insulated from RA, and undergo cell cycle arrest. After birth, male germ cells resume proliferation and undergo a transition to spermatogonia, which are destined to develop into haploid spermatozoa via spermatogenesis. Recent findings indicate that RA levels change periodically in adult testes to direct not only meiotic initiation, but also other key developmental transitions to ensure that spermatogenesis is precisely organized for the prodigious output of sperm. This review focuses on how female and male germ cells develop in the ovary and testis, respectively, and the role of RA in this process.
Assuntos
Oócitos/metabolismo , Ovário/crescimento & desenvolvimento , Espermatozoides/metabolismo , Testículo/crescimento & desenvolvimento , Tretinoína/metabolismo , Animais , Feminino , Gametogênese , Humanos , Masculino , Oócitos/citologia , Ovário/citologia , Ovário/metabolismo , Espermatozoides/citologia , Testículo/citologia , Testículo/metabolismoRESUMO
The mammalian sex chromosomes harbor an abundance of newly acquired ampliconic genes, although their functions require elucidation [1-9]. Here, we demonstrate that the X-linked Slx and Slxl1 ampliconic gene families represent mouse-specific neofunctionalized copies of a meiotic synaptonemal complex protein, Sycp3. In contrast to the meiotic role of Sycp3, CRISPR-loxP-mediated multi-megabase deletions of the Slx (5 Mb) and Slxl1 (2.3Mb) ampliconic regions result in post-meiotic defects, abnormal sperm, and male infertility. Males carrying Slxl1 deletions sire more male offspring, whereas males carrying Slx and Slxl1 duplications sire more female offspring, which directly correlates with Slxl1 gene dosage and gene expression levels. SLX and SLXL1 proteins interact with spindlin protein family members (SPIN1 and SSTY1/2) and males carrying Slxl1 deletions downregulate a sex chromatin modifier, Scml2, leading us to speculate that Slx and Slxl1 function in chromatin regulation. Our study demonstrates how newly acquired X-linked genes can rapidly evolve new and essential functions and how gene amplification can increase sex chromosome transmission.
Assuntos
Fertilidade/genética , Genes Ligados ao Cromossomo X/genética , Família Multigênica/genética , Cromossomos Sexuais/genética , Razão de Masculinidade , Animais , Feminino , Dosagem de Genes , Expressão Gênica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBARESUMO
The ubiquitin proteasome system regulates meiotic recombination in yeast through its association with the synaptonemal complex, a 'zipper'-like structure that holds homologous chromosome pairs in synapsis during meiotic prophase I. In mammals, the proteasome activator subunit PA200 targets acetylated histones for degradation during somatic DNA double strand break repair and during histone replacement during spermiogenesis. We investigated the role of the testis-specific proteasomal subunit α4s (PSMA8) during spermatogenesis, and found that PSMA8 was localized to and dependent on the central region of the synaptonemal complex. Accordingly, synapsis-deficient mice show delocalization of PSMA8. Moreover, though Psma8-deficient mice are proficient in meiotic homologous recombination, there are alterations in the proteostasis of several key meiotic players that, in addition to the known substrate acetylated histones, have been shown by a proteomic approach to interact with PSMA8, such as SYCP3, SYCP1, CDK1 and TRIP13. These alterations lead to an accumulation of spermatocytes in metaphase I and II which either enter massively into apoptosis or give rise to a low number of aberrant round spermatids that apoptose before histone replacement takes place.
Assuntos
Fertilidade/genética , Infertilidade Masculina/genética , Metáfase/genética , Complexo de Endopeptidases do Proteassoma/genética , Subunidades Proteicas/genética , Animais , Apoptose/genética , Modelos Animais de Doenças , Feminino , Masculino , Camundongos , Camundongos Knockout , Proteínas Nucleares/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Subunidades Proteicas/metabolismo , Espermatócitos/metabolismo , Espermatogênese/genética , Complexo Sinaptonêmico/metabolismo , Testículo/citologia , Testículo/metabolismoRESUMO
The germ line provides the cellular link between generations of multicellular organisms, its cells entering the meiotic cell cycle only once each generation. However, the mechanisms governing this initiation of meiosis remain poorly understood. Here, we examined cells undergoing meiotic initiation in mice, and we found that initiation involves the dramatic upregulation of a transcriptional network of thousands of genes whose expression is not limited to meiosis. This broad gene expression program is directly upregulated by STRA8, encoded by a germ cell-specific gene required for meiotic initiation. STRA8 binds its own promoter and those of thousands of other genes, including meiotic prophase genes, factors mediating DNA replication and the G1-S cell-cycle transition, and genes that promote the lengthy prophase unique to meiosis I. We conclude that, in mice, the robust amplification of this extraordinarily broad transcription program by a common factor triggers initiation of meiosis.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Ciclo Celular , Regulação da Expressão Gênica , Células Germinativas/fisiologia , Meiose , Animais , DNA/metabolismo , Redes Reguladoras de Genes , Camundongos , Regiões Promotoras Genéticas , Ligação ProteicaRESUMO
Spermatogenesis has been intensely studied in rodents but remains poorly understood in humans. Here, we used single-cell RNA sequencing to analyze human testes. Clustering analysis of neonatal testes reveals several cell subsets, including cell populations with characteristics of primordial germ cells (PGCs) and spermatogonial stem cells (SSCs). In adult testes, we identify four undifferentiated spermatogonia (SPG) clusters, each of which expresses specific marker genes. We identify protein markers for the most primitive SPG state, allowing us to purify this likely SSC-enriched cell subset. We map the timeline of male germ cell development from PGCs through fetal germ cells to differentiating adult SPG stages. We also define somatic cell subsets in both neonatal and adult testes and trace their developmental trajectories. Our data provide a blueprint of the developing human male germline and supporting somatic cells. The PGC-like and SSC markers are candidates to be used for SSC therapy to treat infertility.
Assuntos
Análise de Célula Única/métodos , Testículo/citologia , Adulto , Diferenciação Celular , Células Cultivadas , Humanos , Recém-Nascido , Masculino , Espermatogônias/citologia , Espermatogônias/metabolismo , Testículo/crescimento & desenvolvimentoRESUMO
Meiotic synapsis and recombination ensure correct homologous segregation and genetic diversity. Asynapsed homologs are transcriptionally inactivated by meiotic silencing, which serves a surveillance function and in males drives meiotic sex chromosome inactivation. Silencing depends on the DNA damage response (DDR) network, but how DDR proteins engage repressive chromatin marks is unknown. We identify the histone H3-lysine-9 methyltransferase SETDB1 as the bridge linking the DDR to silencing in male mice. At the onset of silencing, X chromosome H3K9 trimethylation (H3K9me3) enrichment is downstream of DDR factors. Without Setdb1, the X chromosome accrues DDR proteins but not H3K9me3. Consequently, sex chromosome remodeling and silencing fail, causing germ cell apoptosis. Our data implicate TRIM28 in linking the DDR to SETDB1 and uncover additional factors with putative meiotic XY-silencing functions. Furthermore, we show that SETDB1 imposes timely expression of meiotic and post-meiotic genes. Setdb1 thus unites the DDR network, asynapsis, and meiotic chromosome silencing.
Assuntos
Pareamento Cromossômico , Dano ao DNA , Inativação Gênica , Código das Histonas , Histona-Lisina N-Metiltransferase/metabolismo , Animais , Apoptose , Reparo do DNA , Histona-Lisina N-Metiltransferase/genética , Histonas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteína 28 com Motivo Tripartido/genética , Proteína 28 com Motivo Tripartido/metabolismoRESUMO
Isolating discrete populations of germ cells from the mouse testis is challenging, because the adult testis contains germ cells at every step of spermatogenesis, in addition to somatic cells. We present a novel method for isolating precise, high-purity populations of male germ cells. We first synchronize germ cell development in vivo by manipulating retinoic acid metabolism, and perform histological staging to verify synchronization. We use fluorescence-activated cell sorting to separate the synchronized differentiating germ cells from contaminating somatic cells and undifferentiated spermatogonia. We achieve ~90% purity at each step of development from undifferentiated spermatogonia through late meiotic prophase. Utilizing this "3â¯S" method (synchronize, stage, and sort), we can separate germ cell types that were previously challenging or impossible to distinguish, with sufficient yield for epigenetic and biochemical studies. 3â¯S expands the toolkit of germ cell sorting methods, and should facilitate detailed characterization of molecular and biochemical changes that occur during the mitotic and meiotic phases of spermatogenesis.
Assuntos
Citometria de Fluxo/métodos , Células Germinativas/citologia , Espermatogônias/citologia , Animais , Masculino , Meiose/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Espermatogênese/fisiologia , Testículo/citologia , Tretinoína/metabolismo , Tretinoína/farmacologiaRESUMO
Meiotic cells undergo genetic exchange between homologs through programmed DNA double-strand break (DSB) formation, recombination and synapsis. In mice, the DNA damage-regulated phosphatidylinositol-3-kinase-like kinase (PIKK) ATM regulates all of these processes. However, the meiotic functions of the PIKK ATR have remained elusive, because germline-specific depletion of this kinase is challenging. Here we uncover roles for ATR in male mouse prophase I progression. ATR deletion causes chromosome axis fragmentation and germ cell elimination at mid pachynema. This elimination cannot be rescued by deletion of ATM and the third DNA damage-regulated PIKK, PRKDC, consistent with the existence of a PIKK-independent surveillance mechanism in the mammalian germline. ATR is required for synapsis, in a manner genetically dissociable from DSB formation. ATR also regulates loading of recombinases RAD51 and DMC1 to DSBs and recombination focus dynamics on synapsed and asynapsed chromosomes. Our studies reveal ATR as a critical regulator of mouse meiosis.