Loss of R2D2 Proteins ROPN1 and ROPN1L Causes Defects in Murine Sperm Motility, Phosphorylation, and Fibrous Sheath Integrity
Loss of R2D2 Proteins ROPN1 and ROPN1L Causes Defects in Murine Sperm Motility, Phosphorylation, and Fibrous Sheath Integrity
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DOI：
10.1095/biolreprod.112.105262
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发表时间：
2013-02-01
影响因子：
3.6
通讯作者：
Carr, Daniel W.
中科院分区：
文献类型：
--
作者：
Fiedler, Sarah E.;Dudiki, Tejasvi;Carr, Daniel W.
纤维鞘是精子所特有的一种鞭毛状细胞骨架结构，它包裹着外部致密的纤维和轴丝。它的主要成分是A-激酶锚定蛋白(AKAPs)3和4，这表明FS通过运动所需的信号通路的支架来影响鞭毛跳动。精子蛋白ROPN1和ROPN1L与AKAP3结合。为了确定ROPN1和ROPN1L在精子功能中的作用，我们创建了ROPN1缺陷小鼠(RKO)、ROPN1L缺陷小鼠(RLKO)和双基因敲除小鼠(DKO)。三个品系的小鼠均具有正常的睾丸形态和生精能力。只有DKO在精子形态上有明显的缺陷(主精子变薄和碎裂)，并伴随着AKAP3水平的下降。RLKO小鼠的精子活力略有下降，而ROPN1水平则有所增加。RKO小鼠运动能力中度受损，ROPN1L水平升高。Dko精子不活动。我们之前已经确定RKO雄性小鼠是不育的，而DKO雄性小鼠是不育的。总之，这些数据表明，在缺乏相反蛋白的情况下，ROPN1L和ROPN1相互补偿，可能是为了维持AKAP3在FS中的掺入。缺乏ROPN1L的小鼠精子表现出cAMP依赖的蛋白激酶(PKA)对270 kDa蛋白(可能是FSCB)的磷酸化和获能诱导的酪氨酸磷酸化的减少。缺乏ROPN1的小鼠精子的FSCB水平降低，未获能精子的酪氨酸磷酸化水平增加。这些数据表明，ROPN1和ROPN1L突变可以导致FS完整性、精子活力和依赖PKA的信号传递过程中的缺陷，导致男性不育。
The fibrous sheath (FS) is a flagellar cytoskeletal structure unique to sperm that surrounds the outer dense fibers and axoneme. Its primary components are A-kinase anchoring proteins (AKAPs) 3 and 4, which suggests that the FS affects flagellar beating via the scaffolding of signaling pathways necessary for motility. Sperm proteins ROPN1 and ROPN1L bind AKAP3. To determine the role of ROPN1 and ROPN1L in sperm function, we created mice deficient in ROPN1 (RKO), mice deficient in ROPN1L (RLKO), and double knockout mice (DKO). All three strains of mice had normal testicular morphology and spermatogenesis. Only the DKOs had obvious defects in sperm morphology (thinning and shredding of the principal piece), which was accompanied by a reduction in AKAP3 levels. RLKO mice had slightly reduced sperm motility and increased levels of ROPN1. RKO mice had moderately impaired motility and increased levels of ROPN1L. DKO sperm were immotile. We have previously determined that RKO male mice are subfertile, and DKO males are infertile. Together these data indicate that ROPN1L and ROPN1 compensate for each other in the absence of the opposing protein, possibly to maintain AKAP3 incorporation in the FS. Sperm from mice lacking ROPN1L exhibited reductions in both cAMP-dependent protein kinase (PKA) phosphorylation of a 270-kDa protein (perhaps FSCB), and in capacitation-induced tyrosine phosphorylation. Sperm from mice lacking ROPN1 had reduced levels of FSCB and increased tyrosine phosphorylation of noncapacitated sperm. These data demonstrate that mutations in ROPN1 and ROPN1L can cause defects in FS integrity, sperm motility, and PKA-dependent signaling processes, leading to male infertility.