From the Department of Anatomy and Cell Biology, Philipps-University, 35037 Marburg, Germany,; the Department of Anesthesiology, University Hospital Düsseldorf, 40225 Düsseldorf, Germany, and.
the Department of Biochemistry and Molecular Biology, Mayo Clinic College of Medicine, Rochester, Minnesota 55905.
J Biol Chem. 2011 Mar 11;286(10):7938-7946. doi: 10.1074/jbc.M110.142836. Epub 2010 Dec 27.
Ca(2+) and Ca(2+)-dependent signals are essential for sperm maturation and fertilization. In mouse sperm the plasma membrane Ca(2+)-ATPase (PMCA) isoform 4 plays a crucial role in Ca(2+) transport. The two major splice variants of PMCA4 are PMCA4a and PMCA4b. PMCA4a differs from PMCA4b in the mechanism of calmodulin binding and activation. PMCA4a shows a much higher basal activity and is more effective than PMCA4b in returning Ca(2+) to resting levels. Knock-out mice carrying a PMCA4-null mutation are infertile because their sperm cannot achieve a hyperactivated state of motility. As sperm reach functional maturity during their transit through the epididymis, the expression of PMCA4a and 4b was assessed in bull testis and epididymis. Quantitative PCR revealed that PMCA4b is the major splice variant in testis, caput, and corpus epididymidis. In contrast, PMCA4a is the major splice variant in cauda epididymidis, whereas sperm are transcriptionally silent. Immunohistochemical staining using a new antibody against bovine PMCA4a located the PMCA4a to the apical membrane of the epithelium of cauda epididymidis, whereas testis, caput, and corpus epididymidis were negative. Western blotting of testis, epididymis, and sperm isolated from caput and cauda epididymidis showed a much higher level of PMCA4a in cauda epididymidis and sperm from cauda epididymidis compared with testis membranes and sperm from caput epididymidis. These findings suggest that PMCA4a is transferred to bovine sperm membranes in cauda epididymidis. This isoform switch may facilitate a higher calcium turnover in sperm necessary to traverse the female genital tract.
钙离子和钙离子依赖的信号对于精子成熟和受精至关重要。在小鼠精子中，质膜钙泵（PMCA）同工型 4 在钙转运中起着关键作用。PMCA4 的两个主要剪接变体是 PMCA4a 和 PMCA4b。PMCA4a 在钙调蛋白结合和激活的机制上与 PMCA4b 不同。PMCA4a 具有更高的基础活性，并且比 PMCA4b 更有效地将 Ca2+ 返回到静息水平。携带 PMCA4 缺失突变的敲除小鼠不育，因为它们的精子无法达到运动的超激活状态。由于精子在通过附睾的过程中达到功能成熟，因此评估了公牛睾丸和附睾中 PMCA4a 和 4b 的表达。定量 PCR 显示 PMCA4b 是睾丸、附睾头部和体部的主要剪接变体。相比之下，PMCA4a 是附睾尾部的主要剪接变体，而精子是转录沉默的。使用针对牛 PMCA4a 的新抗体进行的免疫组织化学染色将 PMCA4a 定位到附睾尾部的上皮顶膜，而睾丸、附睾头部和体部均为阴性。从附睾头部和尾部分离的睾丸、附睾和精子的 Western 印迹显示，附睾尾部和精子中的 PMCA4a 水平明显高于睾丸膜和附睾头部的精子。这些发现表明 PMCA4a 在附睾尾部转移到牛精子膜上。这种同工型转换可能促进了精子中更高的钙离子周转率，这对于穿越女性生殖道是必要的。
