Molecular Biology and Genetics Unit, Jawaharlal Nehru Centre for Advanced Scientific Research, Jakkur PO., Bangalore, 560064, India.
Novo Nordisk Foundation Center for Protein Research, Faculty of Health and Medical Sciences, University of Copenhagen, Blegdamsvej 3B, 2200, Copenhagen, Denmark.
Epigenetics Chromatin. 2019 Sep 7;12(1):53. doi: 10.1186/s13072-019-0300-y.
TH2B is a major histone variant that replaces about 80-85% of somatic H2B in mammalian spermatocytes and spermatids. The post-translational modifications (PTMs) on TH2B have been well characterised in spermatocytes and spermatids. However, the biological function(s) of these PTMs on TH2B have not been deciphered in great detail. In our attempt to decipher the unique function(s) of histone variant TH2B, we detected the modification in the N-terminal tail, Serine 11 phosphorylation on TH2B (TH2BS11ph) in spermatocytes.
The current study is aimed at understanding the function of the TH2BS11ph modification in the context of processes that occur during meiotic prophase I. Immunofluorescence studies with the highly specific antibodies revealed that TH2BS11ph histone mark is enriched in the unsynapsed axes of the sex body and is associated with XY body-associated proteins like Scp3, γH2AX, pATM, ATR, etc. Genome-wide occupancy studies as determined by ChIP sequencing experiments in P20 C57BL6 mouse testicular cells revealed that TH2BS11ph is enriched in X and Y chromosomes confirming the immunofluorescence staining pattern in the pachytene spermatocytes. Apart from the localisation of this modification in the XY body, TH2BS11ph is majorly associated with H3K4me3-containing genomic regions like gene promoters, etc. These data were also found to corroborate with the ChIP sequencing data of TH2BS11ph histone mark carried out in P12 C57BL6 mouse testicular cells, wherein we found the predominant localisation of this modification at H3K4me3-containing genomic regions. Mass spectrometry analysis of proteins that associate with TH2BS11ph-containing mononucleosomes revealed key proteins linked with the functions of XY body, pericentric heterochromatin and transcription.
TH2BS11ph modification is densely localised in the unsynapsed axes of the XY body of the pachytene spermatocyte. By ChIP sequencing studies in mouse P12 and P20 testicular cells, we demonstrate that TH2BS11ph is predominantly associated with H3K4me3 positive genomic regions like gene promoters, etc. We propose that TH2BS11ph modification could act alone or in concert with other histone modifications to recruit the appropriate transcription or XY body recombination protein machinery at specific genomic loci.
TH2B 是一种主要的组蛋白变体，在哺乳动物精母细胞和精细胞中取代约 80-85%的体细胞 H2B。TH2B 的翻译后修饰（PTMs）在精母细胞和精细胞中已经得到了很好的描述。然而，这些 TH2B PTM 的生物学功能尚未被详细破译。在我们试图破译组蛋白变体 TH2B 的独特功能的过程中，我们在精母细胞中检测到了 N 端尾巴上的修饰，即 TH2B 的丝氨酸 11 磷酸化（TH2BS11ph）。
本研究旨在了解 TH2BS11ph 修饰在减数分裂前期 I 过程中发生的过程中的功能。使用高度特异性抗体的免疫荧光研究表明，TH2BS11ph 组蛋白标记在性体的未配对轴中富集，并与 XY 体相关蛋白如 Scp3、γH2AX、pATM、ATR 等相关。通过在 P20 C57BL6 小鼠睾丸细胞中进行的 ChIP 测序实验确定的全基因组占据研究表明，TH2BS11ph 富含 X 和 Y 染色体，证实了精母细胞中减数分裂前期的免疫荧光染色模式。除了这种修饰在 XY 体中的定位外，TH2BS11ph 主要与 H3K4me3 包含的基因组区域（如基因启动子等）相关。这些数据还与在 P12 C57BL6 小鼠睾丸细胞中进行的 TH2BS11ph 组蛋白标记的 ChIP 测序数据相吻合，我们发现这种修饰的主要定位在 H3K4me3 包含的基因组区域。与 TH2BS11ph 包含的单核小体相关的蛋白质的质谱分析揭示了与 XY 体、着丝粒异染色质和转录功能相关的关键蛋白质。
TH2BS11ph 修饰在减数分裂前期精母细胞的 XY 体的未配对轴中密集定位。通过在小鼠 P12 和 P20 睾丸细胞中的 ChIP 测序研究，我们证明 TH2BS11ph 主要与 H3K4me3 阳性基因组区域（如基因启动子等）相关。我们提出，TH2BS11ph 修饰可以单独或与其他组蛋白修饰一起，在特定基因组位点募集适当的转录或 XY 体重组蛋白机制。
