Section of Anatomy, Department of Public Health and Cell Biology, University of Rome Tor Vergata, Rome, Italy.
PLoS One. 2012;7(6):e39729. doi: 10.1371/journal.pone.0039729. Epub 2012 Jun 22.
The chromatoid body (CB) is a unique structure of male germ cells composed of thin filaments that condense into a perinuclear organelle after meiosis. Due to the presence of proteins involved in different steps of RNA metabolism and of different classes of RNAs, including microRNAs (miRNAs), the CB has been recently suggested to function as an RNA processing centre. Herein, we show that the RNA binding protein SAM68 transiently localizes in the CB, in concomitance with the meiotic divisions of mouse spermatocytes. Precise staging of the seminiferous tubules and co-localization studies with MVH and MILI, two well recognized CB markers, documented that SAM68 transiently associates with the CB in secondary spermatocytes and early round spermatids. Furthermore, although SAM68 co-immunoprecipitated with MVH in secondary spermatocytes, its ablation did not affect the proper localization of MVH in the CB. On the other hand, ablation of the CB constitutive component MIWI did not impair association of SAM68 with the CB. Isolation of CBs from Sam68 wild type and knockout mouse testes and comparison of their protein content by mass spectrometry indicated that Sam68 ablation did not cause overall alterations in the CB proteome. Lastly, we found that SAM68 interacts with DROSHA and DICER in secondary spermatocytes and early round spermatids and that a subset of miRNAs were altered in Sam68(-/-) germ cells. These results suggest a novel role for SAM68 in the miRNA pathway during spermatogenesis.
染色质体（CB）是一种独特的雄性生殖细胞结构，由在减数分裂后凝聚成核周细胞器的细纤维组成。由于存在涉及 RNA 代谢不同步骤的蛋白质和不同种类的 RNA，包括 microRNAs（miRNAs），因此最近有人提出 CB 作为 RNA 处理中心发挥作用。在此，我们显示 RNA 结合蛋白 SAM68 暂时性地定位于 CB 中，与小鼠精母细胞的减数分裂同时发生。精确分期的精小管和与 MVH 和 MILI 的共定位研究，这两个公认的 CB 标志物，证明 SAM68 在次级精母细胞和早期圆形精子中暂时性地与 CB 相关联。此外，尽管 SAM68 在次级精母细胞中与 MVH 共免疫沉淀，但 SAM68 的缺失并不影响 MVH 在 CB 中的正确定位。另一方面，CB 组成成分 MIWI 的缺失不影响 SAM68 与 CB 的关联。从 Sam68 野生型和敲除小鼠睾丸中分离 CB，并通过质谱比较其蛋白质含量，表明 SAM68 的缺失不会导致 CB 蛋白质组的整体改变。最后，我们发现 SAM68 在次级精母细胞和早期圆形精子中与 DROSHA 和 DICER 相互作用，并且 Sam68(-/-)生殖细胞中的一小部分 miRNA 发生改变。这些结果表明 SAM68 在精子发生过程中的 miRNA 途径中具有新的作用。
