RESUMO
Meiosis pairs and segregates homologous chromosomes and thereby forms haploid germ cells to compensate the genome doubling at fertilization. Homologue pairing in many eukaryotic species depends on formation of DNA double strand breaks (DSBs) during early prophase I when telomeres begin to cluster at the nuclear periphery (bouquet stage). By fluorescence in situ hybridization criteria, we observe that mid-preleptotene and bouquet stage frequencies are altered in male mice deficient for proteins required for recombination, ubiquitin conjugation and telomere length control. The generally low frequencies of mid-preleptotene spermatocytes were significantly increased in male mice lacking recombination proteins SPO11, MEI1, MLH1, KU80, ubiquitin conjugating enzyme HR6B, and in mice with only one copy of the telomere length regulator Terf1. The bouquet stage was significantly enriched in Atm(-/-), Spo11(-/-), Mei1(m1Jcs/m1Jcs), Mlh1(-/-), Terf1(+/-) and Hr6b(-/-) spermatogenesis, but not in mice lacking recombination proteins DMC1 and HOP2, the non-homologous end-joining DNA repair factor KU80 and the ATM downstream effector GADD45a. Mice defective in spermiogenesis (Tnp1(-/-), Gmcl1(-/-), Asm(-/-)) showed wild-type mid-preleptotene and bouquet frequencies. A low frequency of bouquet spermatocytes in Spo11(-/-)Atm(-/-) spermatogenesis suggests that DSBs contribute to the Atm(-/-)-correlated bouquet stage exit defect. Insignificant changes of bouquet frequencies in mice with defects in early stages of DSB repair (Dmc1(-/-), Hop2(-/-)) suggest that there is an ATM-specific influence on bouquet stage duration. Altogether, it appears that several pathways influence telomere dynamics in mammalian meiosis.
Assuntos
Meiose/genética , Mutação , Animais , Proteínas Mutadas de Ataxia Telangiectasia , Proteínas de Ciclo Celular/genética , Quebras de DNA de Cadeia Dupla , Reparo do DNA , Proteínas de Ligação a DNA/deficiência , Proteínas de Ligação a DNA/genética , Endodesoxirribonucleases , Esterases/deficiência , Esterases/genética , Hibridização in Situ Fluorescente , Masculino , Camundongos , Camundongos Knockout , Prófase/genética , Proteínas Serina-Treonina Quinases/deficiência , Proteínas Serina-Treonina Quinases/genética , Recombinação Genética , Espermatócitos/citologia , Espermatócitos/metabolismo , Espermatogênese/genética , Telômero/genética , Proteínas Supressoras de Tumor/deficiência , Proteínas Supressoras de Tumor/genéticaRESUMO
Little is known about the first meiotic prophase stages in the human female because these occur during fetal life, and only a few studies have addressed aneuploid human oocytes. In this paper, the synaptic process in the meiotic prophase in three 47, XX+18 cases is analyzed. A complete study of the dynamics of centromeres and telomeres, cohesin core and synapsis development in aneuploid female meiosis was performed. Investigation of chromosome dynamics in prophase of trisomy 18 oocytes show that these events follow the major patterns seen earlier in euploid oocytes. However, there is a significant delay in the resolution of bouquet topology which could relate to the presence of a surplus chromosome 18 axial element in zygotene oocytes. Pachytene oocytes displayed normal synapsis among the three chromosome 18s. However, in some oocytes the surplus chromosome 18 core was aligned to the bivalent 18. As ataxia telangiectasia and Rad3 related kinase (ATR) has been described as a marker for late-pairing chromosomes in mice, ATR distribution was analyzed in human meiocytes--spermatocytes, euploid oocytes and trisomic oocytes. In contrast to the observations made in mice, no preferential staining for late-pairing chromosomes was observed in humans. In the cases studied, bivalent synapses progressed as in a normal ovary, contrasting with the hypothesis that a surplus chromosome can modify pairing of other chromosomes.
Assuntos
Pareamento Cromossômico , Cromossomos Humanos Par 18 , Prófase Meiótica I , Oócitos/fisiologia , Trissomia , Proteínas Mutadas de Ataxia Telangiectasia , Proteínas de Ciclo Celular/análise , Células Cultivadas , Centrômero , Feminino , Imunofluorescência , Humanos , Processamento de Imagem Assistida por Computador , Hibridização in Situ Fluorescente , Masculino , Microscopia de Fluorescência , Oócitos/química , Diagnóstico Pré-Natal , Proteínas Serina-Treonina Quinases/análise , Espermatócitos/químicaRESUMO
Chromosome segregation errors are a significant cause of aneuploidy among human neonates and often result from errors in female meiosis that occur during fetal life. For the latter reason, little is known about chromosome dynamics during female prophase I. Here, we analyzed chromosome reorganization, and centromere and telomere dynamics in meiosis in the human female by immunofluorescent staining of the SYCP3 and SYCP1 synaptonemal complex proteins and the course of recombinational DNA repair by IF of phospho-histone H2A.X (gamma-H2AX), RPA and MLH1 recombination proteins. We found that SYCP3, but not SYCP1, aggregates appear in the preleptotene nucleus and some persist up to pachytene. Telomere clustering (bouquet stage) in oocytes lasted from late-leptotene to early pachytene-significantly longer than in the male. Leptotene and zygotene oocytes and spermatocytes showed strong gamma-H2AX labeling, while gamma-H2AX patches, which colocalized with RPA, were present on SYCP1-tagged pachytene SCs. This was rarely seen in the male and may suggest that synapsis installs faster with respect to progression of recombinational double-strand break repair or that the latter is slower in the female. It is speculated that the presence of gamma-H2AX into pachytene highlights female-specific peculiarities of recombination, chromosome behavior and checkpoint control that may contribute to female susceptibility for aneuploidy.