Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 111
Filtrar
1.
Cell ; 165(2): 382-95, 2016 04 07.
Artigo em Inglês | MEDLINE | ID: mdl-27040500

RESUMO

Gene duplication is a major evolutionary force driving adaptation and speciation, as it allows for the acquisition of new functions and can augment or diversify existing functions. Here, we report a gene duplication event that yielded another outcome--the generation of antagonistic functions. One product of this duplication event--UPF3B--is critical for the nonsense-mediated RNA decay (NMD) pathway, while its autosomal counterpart--UPF3A--encodes an enigmatic protein previously shown to have trace NMD activity. Using loss-of-function approaches in vitro and in vivo, we discovered that UPF3A acts primarily as a potent NMD inhibitor that stabilizes hundreds of transcripts. Evidence suggests that UPF3A acquired repressor activity through simple impairment of a critical domain, a rapid mechanism that may have been widely used in evolution. Mice conditionally lacking UPF3A exhibit "hyper" NMD and display defects in embryogenesis and gametogenesis. Our results support a model in which UPF3A serves as a molecular rheostat that directs developmental events.


Assuntos
Desenvolvimento Embrionário , Genes Duplicados , Degradação do RNAm Mediada por Códon sem Sentido , Proteínas de Ligação a RNA/metabolismo , Animais , Linhagem Celular Tumoral , Evolução Molecular , Gametogênese , Células HeLa , Humanos , Camundongos
2.
Genes Dev ; 37(13-14): 640-660, 2023 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-37553262

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/genética
3.
Development ; 151(14)2024 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-38884383

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 Helicases
4.
Proc Natl Acad Sci U S A ; 121(9): e2320129121, 2024 Feb 27.
Artigo em Inglês | MEDLINE | ID: mdl-38377195

RESUMO

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 Nuclear
5.
Development ; 150(21)2023 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-37800308

RESUMO

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/metabolismo
6.
Reproduction ; 165(2): 183-196, 2023 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-36395073

RESUMO

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/metabolismo
7.
PLoS Genet ; 16(1): e1008515, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31914128

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/metabolismo
8.
PLoS Genet ; 15(8): e1008316, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31437213

RESUMO

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/metabolismo
9.
Genes Dev ; 27(13): 1484-94, 2013 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-23824539

RESUMO

In mammals, homologs that fail to synapse during meiosis are transcriptionally inactivated. This process, meiotic silencing, drives inactivation of the heterologous XY bivalent in male germ cells (meiotic sex chromosome inactivation [MSCI]) and is thought to act as a meiotic surveillance mechanism. The checkpoint protein ATM and Rad3-related (ATR) localizes to unsynapsed chromosomes, but its role in the initiation and maintenance of meiotic silencing is unknown. Here we show that ATR has multiple roles in silencing. ATR first regulates HORMA (Hop1, Rev7, and Mad2) domain protein HORMAD1/2 phosphorylation and localization of breast cancer I (BRCA1) and ATR cofactors ATR-interacting peptide (ATRIP)/topoisomerase 2-binding protein 1 (TOPBP1) at unsynapsed axes. Later, it acts as an adaptor, transducing signaling at unsynapsed axes into surrounding chromatin in a manner that requires interdependence with mediator of DNA damage checkpoint 1 (MDC1) and H2AFX. Finally, ATR catalyzes histone H2AFX phosphorylation, the epigenetic event leading to gene inactivation. Using a novel genetic strategy in which MSCI is used to silence a chosen gene in pachytene, we show that ATR depletion does not disrupt the maintenance of silencing and that silencing comprises two phases: The first is dynamic and reversible, and the second is stable and irreversible. Our work identifies a role for ATR in the epigenetic regulation of gene expression and presents a new technique for ablating gene function in the germline.


Assuntos
Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Regulação da Expressão Gênica , Inativação Gênica , Meiose , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Animais , Proteínas Mutadas de Ataxia Telangiectasia , Cromossomos/metabolismo , Histonas/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Masculino , Camundongos , Fosforilação , Transporte Proteico/genética , Proteínas Repressoras/metabolismo
10.
Development ; 144(17): 3022-3030, 2017 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-28851723

RESUMO

Spermatogonial stem cells (SSCs) are crucial for maintaining spermatogenesis throughout life, and understanding how these cells function has important implications for understanding male infertility. Recently, various populations of cells harbouring stem cell-like properties have been identified in rodent seminiferous tubules, but deciphering how these cells might fuel spermatogenesis has been difficult, and various models to explain SSC dynamics have been put forward. This Review provides an overview of the organization and timing of spermatogenesis and then discusses these models in light of recent studies of SSC markers, heterogeneity and cell division dynamics, highlighting the evidence for and against each model.


Assuntos
Espermatogônias/citologia , Células-Tronco/citologia , Animais , Biomarcadores/metabolismo , Células Cultivadas , Humanos , Masculino , Espermatogênese , Espermatogônias/metabolismo , Nicho de Células-Tronco , Células-Tronco/metabolismo
11.
Development ; 144(19): 3430-3439, 2017 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-28827392

RESUMO

The human spermatogonial compartment is essential for daily production of millions of sperm. Despite this crucial role, the molecular signature, kinetic behavior and regulation of human spermatogonia are poorly understood. Using human testis biopsies with normal spermatogenesis and by studying marker protein expression, we have identified for the first time different subpopulations of spermatogonia. MAGE-A4 marks all spermatogonia, KIT marks all B spermatogonia and UCLH1 all Apale-dark (Ap-d) spermatogonia. We suggest that at the start of the spermatogenic lineage there are Ap-d spermatogonia that are GFRA1High, likely including the spermatogonial stem cells. Next, UTF1 becomes expressed, cells become quiescent and GFRA1 expression decreases. Finally, GFRA1 expression is lost and subsequently cells differentiate into B spermatogonia, losing UTF1 and acquiring KIT expression. Strikingly, most human Ap-d spermatogonia are out of the cell cycle and even differentiating type B spermatogonial proliferation is restricted. A novel scheme for human spermatogonial development is proposed that will facilitate further research in this field, the understanding of cases of infertility and the development of methods to increase sperm output.


Assuntos
Espermatogônias/citologia , Espermatogônias/metabolismo , Adulto , Idoso , Contagem de Células , Diferenciação Celular , Proliferação de Células , Autorrenovação Celular , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Humanos , Cinética , Masculino , Pessoa de Meia-Idade , Modelos Biológicos , Proteínas Nucleares/metabolismo , Transativadores/metabolismo , Adulto Jovem
12.
Development ; 144(20): 3659-3673, 2017 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-28935708

RESUMO

Spermatogenesis is a dynamic developmental process that includes stem cell proliferation and differentiation, meiotic cell divisions and extreme chromatin condensation. Although studied in mice, the molecular control of human spermatogenesis is largely unknown. Here, we developed a protocol that enables next-generation sequencing of RNA obtained from pools of 500 individually laser-capture microdissected cells of specific germ cell subtypes from fixed human testis samples. Transcriptomic analyses of these successive germ cell subtypes reveals dynamic transcription of over 4000 genes during human spermatogenesis. At the same time, many of the genes encoding for well-established meiotic and post-meiotic proteins are already present in the pre-meiotic phase. Furthermore, we found significant cell type-specific expression of post-transcriptional regulators, including expression of 110 RNA-binding proteins and 137 long non-coding RNAs, most of them previously not linked to spermatogenesis. Together, these data suggest that the transcriptome of precursor cells already contains the genes necessary for cellular differentiation and that timely translation controlled by post-transcriptional regulators is crucial for normal development. These established transcriptomes provide a reference catalog for further detailed studies on human spermatogenesis and spermatogenic failure.


Assuntos
Espermatogênese , Espermatozoides/citologia , Transcriptoma , Adulto , Animais , Biópsia , Diferenciação Celular , Cromatina/química , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Microdissecção e Captura a Laser , Masculino , Meiose , Camundongos , Pessoa de Meia-Idade , Família Multigênica , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/metabolismo , Espermatogônias/citologia , Testículo/citologia
13.
Reproduction ; 159(4): X1, 2020 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-32065737

RESUMO

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.

14.
Proc Natl Acad Sci U S A ; 114(47): E10132-E10141, 2017 11 21.
Artigo em Inglês | MEDLINE | ID: mdl-29109271

RESUMO

Mammalian spermatogenesis is an elaborately organized differentiation process, starting with diploid spermatogonia, which include germ-line stem cells, and ending with haploid spermatozoa. The process involves four pivotal transitions occurring in physical proximity: spermatogonial differentiation, meiotic initiation, initiation of spermatid elongation, and release of spermatozoa. We report how the four transitions are coordinated in mice. Two premeiotic transitions, spermatogonial differentiation and meiotic initiation, were known to be coregulated by an extrinsic signal, retinoic acid (RA). Our chemical manipulations of RA levels in mouse testes now reveal that RA also regulates the two postmeiotic transitions: initiation of spermatid elongation and spermatozoa release. We measured RA concentrations and found that they changed periodically, as also reflected in the expression patterns of an RA-responsive gene, STRA8; RA levels were low before the four transitions, increased when the transitions occurred, and remained elevated thereafter. We found that pachytene spermatocytes, which express an RA-synthesizing enzyme, Aldh1a2, contribute directly and significantly to RA production in testes. Indeed, chemical and genetic depletion of pachytene spermatocytes revealed that RA from pachytene spermatocytes was required for the two postmeiotic transitions, but not for the two premeiotic transitions. We conclude that the premeiotic transitions are coordinated by RA from Sertoli (somatic) cells. Once germ cells enter meiosis, pachytene spermatocytes produce RA to coordinate the two postmeiotic transitions. In combination, these elements underpin the spatiotemporal coordination of spermatogenesis and ensure its prodigious output in adult males.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Aldeído Desidrogenase/genética , Regulação da Expressão Gênica no Desenvolvimento , Espermatogênese/genética , Espermatozoides/metabolismo , Tretinoína/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Aldeído Desidrogenase/metabolismo , Família Aldeído Desidrogenase 1 , Animais , Diferenciação Celular , Masculino , Meiose , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Estágio Paquíteno , Retinal Desidrogenase , Transdução de Sinais , Espermátides/citologia , Espermátides/crescimento & desenvolvimento , Espermátides/metabolismo , Espermatócitos/citologia , Espermatócitos/crescimento & desenvolvimento , Espermatócitos/metabolismo , Espermatogônias/citologia , Espermatogônias/crescimento & desenvolvimento , Espermatogônias/imunologia , Espermatozoides/citologia , Espermatozoides/crescimento & desenvolvimento , Testículo/citologia , Testículo/crescimento & desenvolvimento , Testículo/metabolismo
15.
PLoS Genet ; 13(4): e1006704, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-28380054

RESUMO

The meiosis-specific chromosomal events of homolog pairing, synapsis, and recombination occur over an extended meiotic prophase I that is many times longer than prophase of mitosis. Here we show that, in mice, maintenance of an extended meiotic prophase I requires the gene Meioc, a germ-cell specific factor conserved in most metazoans. In mice, Meioc is expressed in male and female germ cells upon initiation of and throughout meiotic prophase I. Mouse germ cells lacking Meioc initiate meiosis: they undergo pre-meiotic DNA replication, they express proteins involved in synapsis and recombination, and a subset of cells progress as far as the zygotene stage of prophase I. However, cells in early meiotic prophase-as early as the preleptotene stage-proceed to condense their chromosomes and assemble a spindle, as if having progressed to metaphase. Meioc-deficient spermatocytes that have initiated synapsis mis-express CYCLIN A2, which is normally expressed in mitotic spermatogonia, suggesting a failure to properly transition to a meiotic cell cycle program. MEIOC interacts with YTHDC2, and the two proteins pull-down an overlapping set of mitosis-associated transcripts. We conclude that when the meiotic chromosomal program is initiated, Meioc is simultaneously induced so as to extend meiotic prophase. Specifically, MEIOC, together with YTHDC2, promotes a meiotic (as opposed to mitotic) cell cycle program via post-transcriptional control of their target transcripts.


Assuntos
Proteínas de Ciclo Celular/genética , Ciclina A2/biossíntese , Meiose/genética , Prófase/genética , Proteínas de Ligação a RNA/genética , Animais , Proteínas de Ciclo Celular/biossíntese , Pareamento Cromossômico/genética , Ciclina A2/genética , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Camundongos , Mitose/genética , Proteínas de Ligação a RNA/metabolismo , Espermatócitos , Espermatogênese/genética , Espermatogônias/crescimento & desenvolvimento , Espermatogônias/metabolismo
16.
Proc Natl Acad Sci U S A ; 114(47): 12536-12541, 2017 11 21.
Artigo em Inglês | MEDLINE | ID: mdl-29114052

RESUMO

Meiotic synapsis and recombination between homologs permits the formation of cross-overs that are essential for generating chromosomally balanced sperm and eggs. In mammals, surveillance mechanisms eliminate meiotic cells with defective synapsis, thereby minimizing transmission of aneuploidy. One such surveillance mechanism is meiotic silencing, the inactivation of genes located on asynapsed chromosomes, via ATR-dependent serine-139 phosphorylation of histone H2AFX (γH2AFX). Stimulation of ATR activity requires direct interaction with an ATR activation domain (AAD)-containing partner. However, which partner facilitates the meiotic silencing properties of ATR is unknown. Focusing on the best-characterized example of meiotic silencing, meiotic sex chromosome inactivation, we reveal this AAD-containing partner to be the DNA damage and checkpoint protein TOPBP1. Conditional TOPBP1 deletion during pachynema causes germ cell elimination associated with defective X chromosome gene silencing and sex chromosome condensation. TOPBP1 is essential for localization to the X chromosome of silencing "sensors," including BRCA1, and effectors, including ATR, γH2AFX, and canonical repressive histone marks. We present evidence that persistent DNA double-strand breaks act as silencing initiation sites. Our study identifies TOPBP1 as a critical factor in meiotic sex chromosome silencing.


Assuntos
Proteínas de Transporte/genética , Quebras de DNA de Cadeia Dupla , Cromossomos Sexuais/química , Espermatogênese/genética , Inativação do Cromossomo X , Animais , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Proteína BRCA1 , Proteínas de Transporte/metabolismo , Pareamento Cromossômico , Histonas/genética , Histonas/metabolismo , Masculino , Camundongos , Camundongos Knockout , Cromossomos Sexuais/metabolismo , Espermátides/citologia , Espermátides/crescimento & desenvolvimento , Espermátides/metabolismo , Espermatócitos/citologia , Espermatócitos/crescimento & desenvolvimento , Espermatócitos/metabolismo , Espermatogônias/citologia , Espermatogônias/crescimento & desenvolvimento , Espermatogônias/metabolismo , Espermatozoides/citologia , Espermatozoides/crescimento & desenvolvimento , Espermatozoides/metabolismo , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo
17.
Dev Biol ; 443(1): 19-34, 2018 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-30149006

RESUMO

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 "3 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. 3 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/farmacologia
18.
Hum Mol Genet ; 25(24): 5300-5310, 2016 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-27742779

RESUMO

During spermatogenesis, germ cells that fail to synapse their chromosomes or fail to undergo meiotic sex chromosome inactivation (MSCI) are eliminated via apoptosis during mid-pachytene. Previous work showed that Y-linked genes Zfy1 and Zfy2 act as 'executioners' for this checkpoint, and that wrongful expression of either gene during pachytene triggers germ cell death. Here, we show that in mice, Zfy genes are also necessary for efficient MSCI and the sex chromosomes are not correctly silenced in Zfy-deficient spermatocytes. This unexpectedly reveals a triple role for Zfy at the mid-pachytene checkpoint in which Zfy genes first promote MSCI, then monitor its progress (since if MSCI is achieved, Zfy genes will be silenced), and finally execute cells with MSCI failure. This potentially constitutes a negative feedback loop governing this critical checkpoint mechanism.


Assuntos
Proteínas de Ligação a DNA/genética , Espermatócitos/metabolismo , Fatores de Transcrição/genética , Inativação do Cromossomo X/genética , Animais , Masculino , Meiose/genética , Camundongos , Espermatócitos/crescimento & desenvolvimento , Espermatogênese/genética , Cromossomo X/genética
19.
Biol Reprod ; 99(1): 75-86, 2018 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-29590307

RESUMO

This review focuses on those mouse mutations that cause an effect on the morphology, viability, and/or behavior of primordial germ cells (PGCs) and gonocytes at specific steps of their fetal development up to the start of spermatogenesis, a few days after birth. To restrict the area covered, mice with mutations that cause abnormal hormone levels or mutations of genes not expressed in germ cells that secondarily cause spermatogenic problems are not discussed. To make our literature search as comprehensive as possible, Pubmed was searched for "(primordial germ cells OR prospermatogonia OR prespermatogonia OR gonocytes OR spermatogonia or meiosis or spermiogenesis or spermatogenesis) AND mouse AND (knockout or mutant or transgenic)." This search started at 2003 as mutants created earlier were already retrieved for a previous review. The resulting citations were then further selected for complete or partial arrests at the level of PGCs and/or gonocytes. Fifty-nine protein coding genes and two miRNA coding genes were found that arrest the development of PGCs and gonocytes at specific steps providing a better insight into the regulation of the development of these cells. As to be expected, often problems in fetal germ cell development have an effect on the fertility of the mice at adulthood.


Assuntos
Pontos de Checagem do Ciclo Celular/genética , Mutação , Oogênese/genética , Oogônios/citologia , Espermatogênese/genética , Espermatogônias/citologia , Animais , Masculino , Camundongos
20.
Nucleic Acids Res ; 44(20): 9681-9697, 2016 Nov 16.
Artigo em Inglês | MEDLINE | ID: mdl-27431324

RESUMO

Meiotic recombination is essential for fertility in most sexually reproducing species, but the molecular mechanisms underlying this process remain poorly understood in mammals. Here, we show that RNF20-mediated H2B ubiquitination is required for meiotic recombination. A germ cell-specific knockout of the H2B ubiquitination E3 ligase RNF20 results in complete male infertility. The Stra8-Rnf20-/- spermatocytes arrest at the pachytene stage because of impaired programmed double-strand break (DSB) repair. Further investigations reveal that the depletion of RNF20 in the germ cells affects chromatin relaxation, thus preventing programmed DSB repair factors from being recruited to proper positions on the chromatin. The gametogenetic defects of the H2B ubiquitination deficient cells could be partially rescued by forced chromatin relaxation. Taken together, our results demonstrate that RNF20/Bre1p-mediated H2B ubiquitination regulates meiotic recombination by promoting chromatin relaxation, and suggest an old drug may provide a new way to treat some oligo- or azoospermia patients with chromatin relaxation disorders.


Assuntos
Montagem e Desmontagem da Cromatina , Cromatina/genética , Cromatina/metabolismo , Histonas/metabolismo , Meiose , Recombinação Genética , Proteínas Adaptadoras de Transdução de Sinal/deficiência , Animais , Quebras de DNA de Cadeia Dupla , Reparo do DNA , Feminino , Células Germinativas/metabolismo , Infertilidade Masculina/genética , Masculino , Camundongos , Camundongos Knockout , Estágio Paquíteno/genética , Espermatócitos/metabolismo , Espermatogênese/genética , Ubiquitina-Proteína Ligases/deficiência , Ubiquitinação
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA