Department of Pathology and Immunology, Baylor College of Medicine, Houston, Texas, United States of America.
PLoS One. 2011 Feb 25;6(2):e17066. doi: 10.1371/journal.pone.0017066.
Intercellular bridges are evolutionarily conserved structures that connect differentiating germ cells. We previously reported the identification of TEX14 as the first essential intercellular bridge protein, the demonstration that intercellular bridges are required for male fertility, and the finding that intercellular bridges utilize components of the cytokinesis machinery to form. Herein, we report the identification of RNA binding motif protein 44 (RBM44) as a novel germ cell intercellular bridge protein. RBM44 was identified by proteomic analysis after intercellular bridge enrichment using TEX14 as a marker protein. RBM44 is highly conserved between mouse and human and contains an RNA recognition motif of unknown function. RBM44 mRNA is enriched in testis, and immunofluorescence confirms that RBM44 is an intercellular bridge component. However, RBM44 only partially localizes to TEX14-positive intercellular bridges. RBM44 is expressed most highly in pachytene and secondary spermatocytes, but disappears abruptly in spermatids. We discovered that RBM44 interacts with itself and TEX14 using yeast two-hybrid, mammalian two-hybrid, and immunoprecipitation. To define the in vivo function of RBM44, we generated a targeted deletion of Rbm44 in mice. Rbm44 null male mice produce somewhat increased sperm, and show enhanced fertility of unknown etiology. Thus, although RBM44 localizes to intercellular bridges during meiosis, RBM44 is not required for fertility in contrast to TEX14.
细胞间桥是连接分化生殖细胞的进化保守结构。我们之前曾报道过 TEX14 是第一个必需的细胞间桥蛋白，证明细胞间桥对于雄性生育力是必需的，并且发现细胞间桥利用胞质分裂机制的成分形成。在此，我们报告了 RNA 结合基序蛋白 44（RBM44）作为一种新型的生殖细胞细胞间桥蛋白的鉴定。RBM44 是通过使用 TEX14 作为标记蛋白进行细胞间桥富集的蛋白质组学分析鉴定的。RBM44 在小鼠和人类之间高度保守，并且含有一个未知功能的 RNA 识别基序。RBM44 mRNA 在睾丸中富集，免疫荧光证实 RBM44 是细胞间桥的组成部分。然而，RBM44 仅部分定位于 TEX14 阳性细胞间桥。RBM44 在粗线期和次级精母细胞中表达最高，但在精细胞中突然消失。我们发现 RBM44 使用酵母双杂交、哺乳动物双杂交和免疫沉淀与自身和 TEX14 相互作用。为了定义 RBM44 的体内功能，我们在小鼠中生成了 Rbm44 的靶向缺失。Rbm44 缺失的雄性小鼠产生的精子略有增加，并且具有未知病因的生育力增强。因此，尽管 RBM44 在减数分裂期间定位于细胞间桥，但与 TEX14 不同，RBM44 对于生育力不是必需的。
