Regulation of sperm motility by the Rho signaling pathway

Rho 信号通路对精子活力的调节

• 批准号: 8238592
• 负责人: DANIEL W CARR
• 金额: $ 6.3万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-28 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8238592
• 关键词: A kinase anchoring protein; Accounting; Adaptor Signaling Protein; Affect; Binding; Biochemical; Cilia; Couples; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Diagnosis; Dimerization; Disease; Docking; Drug Design; Enzymes; Event; Fertility; Flagella; Genomics; Goals; Human; In Vitro; Infertility; Knockout Mice; Knowledge; Literature; Male Infertility; Measures; Microtubules; Molecular; Mus; Muscle Contraction; Mutant Strains Mice; Names; Pathway interactions; Peptides; Phosphorylation; Physiological; Plant Roots; Process; Protein Binding; Protein Isoforms; Protein phosphatase; Proteins; Publishing; Regulation; Role; Scaffolding Protein; Signal Pathway; Signal Transduction; Slide; Sperm Motility; Testing; Testis; Therapeutic; Ursidae Family; Yeasts; cell motility; in vivo; male; men; protein expression; research study; rho; rho GTP-Binding Proteins; rhophilin; sperm cell; therapy development; yeast two hybrid system

DESCRIPTION (provided by applicant): Suboptimal sperm motility accounts for nearly 30% of all human infertility. As such, a better understanding of the biochemical mechanisms regulating sperm motility may provide therapeutic options for infertile couples. We have recently identified a gene product (ROPN1) that appears to be critical for both sperm motility and fertility. ROPN1 is highly expressed in testis and sperm, and mice lacking ROPN1 have impaired sperm motility and are subfertile or infertile. The expression level of ROPN1 is significantly lower in asthenozoospermic men than in normozoospermic controls. ROPN1 was originally identified as a protein that binds to rhophilin, a binding partner of Rho. Using a yeast two-hybrid analysis, we discovered that ROPN1 also binds to sperm A-kinase anchoring proteins (AKAPs). ROPN1 contains a dimerization/docking domain similar to the regulatory subunit of protein kinase A (PKA). However, outside the docking domain, ROPN1 bears little or no similarity to PKA suggesting they have distinct functions. Two observations suggest that ROPN1 is required for flagellar motility: 1) adding peptides (Ht31) that disrupt the interaction between AKAPs and ROPN1 results in altered protein phosphatase 1 (PP1) activity, GSK3 phosphorylation and arrested sperm motility, and 2) sperm from mutant mice lacking ROPN1 (Ropn1-/-) have reduced motility. Thus, ROPN1 appears to be a key molecule located at the intersection between the cAMP and Rho pathways. Although many articles have been published on the role of cAMP/PKA in sperm, the literature on Rho in sperm is relatively sparse. Inactivation of Rho-GTPase has been shown to reduce mammalian sperm motility and we have recently shown that sperm contain all the protein components of the Rho signaling pathway. Inhibition of PP1, a downstream effector in the Rho pathway, dramatically alters sperm motility. However, to our knowledge, no one has measured the activity of Rho-GTPase from sperm. The goal of this project is to determine if Rho is a critical component of the biochemical machinery regulating sperm motility and if ROPN1 functions as part of this pathway. Successful completion of this proposal will enhance our knowledge of this important regulatory mechanism and thus may facilitate development of treatments for male infertility. PUBLIC HEALTH RELEVANCE: The goal of this proposal is to enhance knowledge of signaling pathways that regulate flagellar function, which is essential to defining normal male fertility. This increased understanding of normal processes should allow for more precise diagnoses of the root cause(s) of infertility, and will also provide more targets for treatment and rational drug design. Commonalities between flagella and cilia in function, protein expression and signaling pathways suggest that these findings may also have wider implications in diagnosis and treatment of ciliary diseases.

描述（由申请人提供）：次优精子活力占所有人类不育症的近30%。因此，更好地了解调节精子活力的生化机制可能为不育夫妇提供治疗选择。我们最近发现了一种基因产物（ROPN 1），它似乎对精子活力和生育力都至关重要。ROPN 1在睾丸和精子中高度表达，缺乏ROPN 1的小鼠精子活力受损，生育力低下或不育。弱精子症患者ROPN 1的表达水平明显低于正常精子症患者。ROPN 1最初被鉴定为与Rho的结合伴侣rhophilin结合的蛋白质。使用酵母双杂交分析，我们发现ROPN 1也结合精子A激酶锚定蛋白（AKAP）。ROPN 1含有一个类似于蛋白激酶A（PKA）调节亚基的二聚/对接结构域。然而，在对接结构域之外，ROPN 1与PKA几乎没有相似性，这表明它们具有不同的功能。两个观察结果表明，ROPN 1是鞭毛运动所必需的：1）添加破坏AKAP和ROPN 1之间相互作用的肽（Ht 31），导致蛋白磷酸酶1（PP 1）活性改变，GSK 3磷酸化和精子运动性停滞，2）缺乏ROPN 1（Ropn 1-/-）的突变小鼠精子运动性降低。因此，ROPN 1似乎是位于cAMP和Rho途径之间的交叉点的关键分子。虽然已经发表了许多关于cAMP/PKA在精子中的作用的文章，但关于精子中Rho的文献相对较少。Rho-GTdR的失活已被证明会降低哺乳动物精子的活力，我们最近发现精子含有Rho信号通路的所有蛋白质组分。PP 1是Rho通路的下游效应子，抑制PP 1可显著改变精子活力。然而，据我们所知，还没有人从精子中测量Rho-GTdR的活性。该项目的目标是确定Rho是否是调节精子活力的生化机制的关键组成部分，以及ROPN 1是否作为该途径的一部分发挥作用。成功完成这项提案将提高我们对这一重要调节机制的认识，从而可能促进男性不育治疗的发展。 公共卫生关系：该提案的目标是提高对调节鞭毛功能的信号通路的认识，这对于定义正常的男性生育力至关重要。这种对正常过程的理解的增加应该允许对不孕症的根本原因进行更精确的诊断，并且还将为治疗和合理的药物设计提供更多的靶点。鞭毛和纤毛在功能、蛋白表达和信号通路方面的共同点表明，这些发现也可能在纤毛疾病的诊断和治疗中具有更广泛的意义。