Establishment and Analysis of Cylicin1 and Cylicin2 deficient mice – a tool to study male factor infertility

cyclin基因家族，cyclin 1 （Cylc1）和cyclin 2 （Cylc2）被发现只在睾丸中表达。CYLC1存在于人类和牛精子的花萼区域，呈近赤道带状和漏斗状结构。cyclin蛋白具有重复的、富含赖氨酸的三肽的特征，这导致蛋白质具有10.55的等电点的整体正电荷。最常见的是，三肽由赖氨酸二肽和天冬氨酸或谷氨酸（KKD/KKE）组成。此外，CYLC1含有一个富含脯氨酸的c端结构域。蛋白质的中心部分由几个重复单元组成，预计它们会形成短的α-螺旋。细胞周期蛋白的功能在很大程度上仍然未知。然而，由于在男性结构异常的圆头精子中发现了核周卵泡形成缺陷，因此讨论了精子发生过程中精子头部形成的作用。猪CYLC2进一步被证明是一种肌动蛋白结合蛋白，可能在圆形精子顶体区域和成熟精子顶体后区域起细胞骨架调节作用。最近，一些不孕症男性被诊断为CYLC1功能突变缺失，这表明CYLC1可能对精子发生和男性生育能力至关重要。该项目旨在进一步全面了解cyclin 1和/或cyclin 2比率丧失的后果。我们设想建立一个男性因素不育的模型。•建立并验证Cylicin 1和Cylicin 2缺失小鼠系。•分析Cylicin 1和Cylicin 2缺失的影响。•将Cyclicin 1突变与Cyclicin 2突变杂交，分析其对精子和生育能力的冗余和补偿效应。•使用蛋白质组学分析任何一种蛋白质损失的分子效应分析将有助于理解细胞周期素的基本生物学，并将作为关于人类数据的原则证明。我们将能够生成细胞周期素缺乏男性因素不育的小鼠模型，并能够分析由缺乏引起的缺陷。最终，本文提出的研究将为更好地了解男性不孕症和帮助制定治疗方案奠定基础。

The family of Cylicin-genes, Cylicin 1 (Cylc1) and Cylicin 2 (Cylc2) are found exclusively expressed in the testis. CYLC1 is detected in the calyx region of human and bovine sperm and presents as a near-equatorial belt-like and funnel shaped structure. Cylicin proteins are characterized by repetitive, lysine-rich tripeptides, which result in an overall positive charge of the protein with an isoelectric point of 10.55. Most frequent, tripeptides consist of lysine dipeptides followed by aspartic or glutamic acid (KKD/KKE). Further, CYLC1 contains a proline-rich C-terminal domain. The central part of the protein comprises several repeating units, which are predicted to form short α-helices. The function of Cylicins remains largely unknown. However, a role in sperm head shaping during spermiogenesis is discussed, since defects in perinuclear theca formation have been identified in structurally abnormal, round-headed sperm in men. Boar CYLC2 was further shown to function as an actin-binding protein, which might serve as a cytoskeletal regulator in the acrosomal region of round spermatids and the postacrosomal region of mature sperm. Recently, several infertile men were diagnosed to harbor loss of function mutations in CYLC1 indicating that Cylcins could be essential for spermatogenesis and male fertility.The project aims at further understanding the consequences of loss of Cylicin 1 and/or Cylicin 2 ratio in a comprehensive matter. We envision to develop a model of male factor infertility.• To establish and validate Cylicin 1 and Cylicin 2 deficient mouse lines.• To analyze the effects of loss of Cylicin 1 and Cylicin 2.• To breed the Cyclicin 1 mutation with the Cylicin 2 mutation to analyze redundant and compensatory effects on sperm and fertility.• To analyze the molecular effects of loss of either protein using proteomicsAnalyses will help to understand the basic biology of Cylicins and will serve as a proof of principle with regard to the human data. We will be able to generate a murine model for Cylicin-deficient male factor infertility and will be able to analyze the defects originating from the deficiency. Ultimately, the research proposed here will lay the basis to better understand infertility in man and help in development of treatment regimen.