睾丸优势表达蛋白FBXO47在减数分裂过程中的作用和机制研究

Spermatocyte arrest was detected in 12% of azoospermia which is caused by meiosis defect. Recent study revealed that post-translational modification (PTM) plays a unique role in regulation of meiosis including ubiquitination. It is still unclear how many proteins are involved in ubiquitination during meiosis and which protein is regulated by ubiquitination. In our human testicular proteomics research, we discovered a series of F-box domain proteins, which could binding with SKP1 and form a SCF complex. SCF complex could play a role of E3 ligase to regulate ubiquitination. In this study, we focus on one of F-box proteins, FBXO47 (F-box protein 47), which is functional unknown. We demonstrate that FBXO47 is predominantly expressed in testis, and only expressed in spermatocyte; in mouse model, FBXO47 deficiency cause meiosis arrest in spermatocyte and male infertility, but not in female. We will access which meiotic event FBXO47 is involved in; discover which meiotic event is defeated in FBXO47 knockout mice; find substrates of SCF-FBXO47 complex E3 ligase. Our study could provide an overall view of FBXO47 in regulation of meiosis.

临床上约有12%的无精子症病人存在减数分裂的异常。随着研究的深入，发现蛋白质翻译后修饰在减数分裂中起到了重要的调控作用。蛋白质泛素化可能作为潜在的重要调控形式参与减数分裂。在我们前期人类睾丸蛋白质组学谱系的研究中，其中有一系列蛋白质具有F-box结构域的蛋白质，该结构域可以结合SKP1，形成SCF复合体E3泛素连接酶调控蛋白质泛素化。这里我们关注到其中一个功能未知蛋白FBXO47。我们前期研究发现小鼠FBXO47在睾丸优势表达，进一步分析发现FBXO47主要表达于初级精母细胞中；缺失了FBXO47的小鼠，雄性不育，雌性可育，生精细胞阻滞在精母细胞减数分裂阶段。本课题拟通过FBXO47定位研究和基因敲除小鼠表型分析揭示FBXO47缺失影响减数分裂的具体环节；筛选和鉴定FBXO47参与SCF复合体E3泛素连接酶的底物，全面阐明FBXO47在减数分裂调控中的作用。

减数分裂障碍常常导致不孕不育的严重结局。在减数分裂过程中，端粒与核膜的锚定是同源染色体配对和重组的必要条件。在这里，我们制备了一种针对F-box蛋白47（FBXO47）的抗体，并鉴定了FBXO47作为端粒蛋白复合物（shelterin）的调节因子，该复合物在第一次减数分裂前期特异表达。FBXO47基因敲除导致雄性不育。我们发现Fbxo47缺陷精母细胞不能形成完整的联会复合体。FBXO47与TRF1/2相互作用，FBXO47的缺失破坏了TRF2的稳定性，导致端粒核膜锚定不稳定，并出现过花束期阻滞。我们的发现揭示了FBXO47在减数分裂过程中稳定端粒shelterin亚单位的新机制。同时，也为男性不育的诊疗提供了新的理论基础。

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