RESUMO
The synaptonemal complex (SC) is the central key structure for meiosis in organisms undergoing sexual reproduction. During meiotic prophase I, homologous chromosomes exchange genetic information at the time they are attached to the lateral elements by specific DNA sequences. Most of these sequences, so far identified, consist of repeat DNA, which are subject to chromatin structural changes during meiotic prophase I. In this work, we addressed the effect of altering the chromatin structure of repeat DNA sequences mediating anchorage to the lateral elements of the SC. Administration of the histone deacetylase inhibitor trichostatin A into live rats caused death of cells in the pachytene stage as well as changes in histone marks along the synaptonemal complex. The most notable effect was partial loss of histone H3 lysine 27 trimethylation. Our work describes the epigenetic landscape of lateral element-associated chromatin and reveals a critical role of histone marks in synaptonemal complex integrity.
Assuntos
Histonas/metabolismo , Sequências Repetitivas de Ácido Nucleico , Complexo Sinaptonêmico/química , Animais , Cromatina/genética , Cromatina/metabolismo , Cromossomos/genética , Cromossomos/metabolismo , Histonas/genética , Masculino , Prófase Meiótica I , Estabilidade Proteica , Ratos , Ratos Wistar , Complexo Sinaptonêmico/genética , Complexo Sinaptonêmico/metabolismo , Testículo/química , Testículo/citologia , Testículo/metabolismoRESUMO
BACKGROUND: The mechanisms of meiotic arrest in human spermatogenesis are poorly known. METHODS AND RESULTS: A testicular biopsy from an azoospermic male showed complete spermatogenesis arrest at the spermatocyte stage, asynapsis, lack of formation of the XY body, partial reversion to a mitotic-like division and cell degeneration both at the prophase and at the abnormal cell divisions. Synaptonemal complex analysis showed minor segments of synapsis and mainly single axes. Fluorescent immunolocalization of meiotic proteins showed normal SYCP3, scarcity of SYCP1, null MLH1 foci, about 10 patches of gamma-H2AX, abnormal presence of BRCA1 among autosomal axes, absence of RAD51 in early and advanced spermatocytes and permanence of gamma-H2AX labelling up to the abnormal spermatocyte divisions that are the most advanced stage reached. There are at least six dominions of evenly packed chromatin resembling that of the normal XY body, but no true XY body. CONCLUSIONS: The protein phenotype and the fine structure of the nuclei are compatible with a deficiency of the processing of double-strand DNA breaks in the zygotene-like spermatocytes, but the features of this defect do not agree with Spo11, Sycp1, Atm and Dmc1 null mutations, which give absence of XY body, synapsis disturbances and spermatocyte apoptosis in mice.
Assuntos
Dano ao DNA , Reparo do DNA , Oligospermia/metabolismo , Espermatócitos/ultraestrutura , Espermatogênese , Complexo Sinaptonêmico/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Adenosina Trifosfatases/genética , Adulto , Proteínas Mutadas de Ataxia Telangiectasia , Proteína BRCA1/análise , Proteínas de Transporte , Proteínas de Ciclo Celular/análise , Proteínas de Ciclo Celular/genética , Núcleo Celular/química , Núcleo Celular/metabolismo , Núcleo Celular/ultraestrutura , Pareamento Cromossômico/genética , DNA/metabolismo , Dano ao DNA/genética , Proteínas de Ligação a DNA/análise , Proteínas de Ligação a DNA/genética , Endodesoxirribonucleases , Esterases/genética , Histonas/análise , Humanos , Masculino , Meiose/genética , Proteína 1 Homóloga a MutL , Mutação , Proteínas Nucleares/análise , Proteínas Nucleares/genética , Oligospermia/genética , Proteínas Serina-Treonina Quinases/análise , Rad51 Recombinase/análise , Espermatócitos/química , Espermatócitos/metabolismo , Espermatogênese/genética , Complexo Sinaptonêmico/química , Complexo Sinaptonêmico/genética , Testículo/patologia , Proteínas Supressoras de Tumor/análiseRESUMO
The formation of the XY body involves the compaction of the extended chromatin to form a mesh of fibrogranular structures. During this process the ribonucleoprotein particles (RNP), which were associated with the chromatin filaments progressively disappear. High resolution immunolocalization indicates that the mature XY body does not contain RNA polymerase II, hnRNPs, or snURNPs. Occasionally chromatin fibrils extend outside of the XY body. These fibrils are frequently associated with nascent RNP fibrils and granules indicating that not all the DNA of the sex chromosomes is transcriptionally inactive. However, transcription is located outside the sex body. The recombination protein Dmc1 is present in nodules associated with the unpaired chromosomal axes of the sex chromosomes located in the XY body. Cytochemical staining methods and in situ hybridization at electron microscopic level show that RNA is present in the unpaired chromosomal axes suggesting that the presence of RNA in the chromosomal axes and in forming synaptonemal complexes is related with the process of final pairing. The sex body and the nucleoli associated with it do not interweave and do not exchange RNA or DNA-containing filaments. These observations indicate that the spatial relation between these structures is just a close proximity, which is, however, very frequent.
Assuntos
Imuno-Histoquímica/métodos , Cromossomos Sexuais/ultraestrutura , Espermatócitos/ultraestrutura , Complexo Sinaptonêmico/ultraestrutura , Animais , Cobaias , Hibridização In Situ , Masculino , RNA/análise , Ratos , Cromossomos Sexuais/química , Espermatócitos/química , Complexo Sinaptonêmico/química , Testículo/citologia , Translocação GenéticaRESUMO
The frequency and distribution of the mismatch repair protein MLH1 was analyzed on synaptonemal complex spreads of chicken oocytes using indirect immunofluorescence. MLH1 foci appeared in late zygotene and their number remains constant throughout pachytene. The average number of foci on autosomal synaptonemal complexes (65.02 +/- 4.02) is in agreement with the number of chiasmata estimated from lampbrush chromosomes. The distribution of foci along the synaptonemal complexes is shown to be nonrandom and nonuniform in terms of the distances between them. It is concluded that MLH1 foci are good markers of crossing over in bird (chicken) meiocytes.