Signal Transduction Laboratory, Centre for Cell Biology, Biology Department, University of Aveiro, 3810-193 Aveiro, Portugal.
Biochem Pharmacol. 2011 Nov 15;82(10):1403-15. doi: 10.1016/j.bcp.2011.02.018. Epub 2011 Mar 5.
Protein phosphorylation is a critical regulatory mechanism in cellular signalling. To this end, PP1 is a major eukaryotic serine/threonine-specific phosphatase whose cellular functions, in turn, depend on complexes it forms with PP1 interacting proteins-PIPs. The importance of the testis/sperm-enriched variant, PP1γ2, in sperm motility and spermatogenesis has previously been shown. Given the key role of PIPs, it is imperative to identify the physiologically relevant PIPs in testis and sperm. Hence, we performed Yeast Two-Hybrid screens of a human testis cDNA library using as baits the different PP1 isoforms and also a proteomic approach aimed at identifying PP1γ2 binding proteins. To the best of our knowledge this is the largest data set of the human testis PP1 interactome. We report the identification of 77 proteins in human testis and 7 proteins in human sperm that bind PP1. The data obtained increased the known PP1 interactome by reporting 72 novel interactions. Confirmation of the interaction of PP1 with 5 different proteins was also further validated by co-immunoprecipitation or protein overlays. The data here presented provides important insights towards the function of these proteins and opens new possibilities for future research. In fact, such diversity in PP1 regulators makes them excellent targets for pharmacological intervention.
蛋白质磷酸化是细胞信号转导的一种关键调节机制。为此，PP1 是一种主要的真核丝氨酸/苏氨酸特异性磷酸酶，其细胞功能又依赖于它与 PP1 相互作用蛋白-PIPs 形成的复合物。先前已经表明，富含睾丸/精子的变体 PP1γ2 在精子运动和精子发生中具有重要作用。鉴于 PIPs 的重要性，鉴定睾丸和精子中生理相关的 PIPs 至关重要。因此，我们使用不同的 PP1 同工型作为诱饵，对人睾丸 cDNA 文库进行了酵母双杂交筛选，还采用了一种蛋白质组学方法来鉴定 PP1γ2 结合蛋白。据我们所知，这是人类睾丸 PP1 相互作用组的最大数据集。我们报告了在人类睾丸中鉴定出 77 种蛋白质和人类精子中鉴定出 7 种与 PP1 结合的蛋白质。所获得的数据通过报告 72 个新的相互作用，增加了已知的 PP1 相互作用组。还通过共免疫沉淀或蛋白质重叠进一步验证了 PP1 与 5 种不同蛋白质相互作用的确认。这里提供的数据为这些蛋白质的功能提供了重要的见解，并为未来的研究开辟了新的可能性。事实上，PP1 调节剂的这种多样性使它们成为药物干预的理想靶点。
