Plasma membrane Ca2+-ATPase 4 in murine epididymis: Secretion of splice variants in the luminal fluid and a role in sperm maturation

Ramkrishna Patel, Amal A. Al-Dossary, Deborah L. Stabley, Carol Barone, Deni S. Galileo, Emanuel E. Strehler, Patricia A. Martin-DeLeon

Research output: Contribution to journalArticle

23 Scopus citations

Abstract

Plasma membrane Ca2+-ATPase isoform 4 (PMCA4) is the primary Ca2+ efflux pump in murine sperm, where it regulates motility. In Pmca4 null sperm, motility loss results in infertility. We have shown that murine sperm PMCA4b interacts with Ca2+/ CaM-dependent serine kinase (CASK) in regulating Ca2+ homeostasis and motility. However, recent work indicated that the bovine PMCA4a splice variant (missing in testis) is epididymally expressed, along with 4b, and may be transferred to sperm. Here we show, via conventional and in situ RT-PCR, that both the splice variants of Pmca4 mRNA are expressed in murine testis and throughout the epididymis. Immunofluorescence localized PMCA4a to the apical membrane of the epididymal epithelium, and Western analysis not only confirmed its presence but showed for the first time that PMCA4a and PMCA4b are secreted in the epididymal luminal fluid (ELF), from which epididymosomes containing PMCA4a were isolated. Flow cytometry indicated the presence of PMCA4a on mature caudal sperm where it was increased ;5-fold compared to caput sperm (detected by Western blotting) and ;2-fold after incubation in ELF, revealing in vitro uptake and implicating PMCA4a in epididymal sperm maturation. Coimmunoprecipitation using pan-PMCA4 antibodies, revealed that both variants associate with CASK, suggesting their presence in a complex. Because they have different kinetic properties for Ca2+ transport and different abilities to bind to CASK, our study suggests a mechanism for combining the functional attributes of both PMCA4 variants, leading to heightened efficiency of the pump in the maintenance of Ca2+ homeostasis, which is crucial for normal motility and male fertility.

Original languageEnglish (US)
Article number6
JournalBiology of Reproduction
Volume89
Issue number1
DOIs
StatePublished - 2013

Keywords

  • CASK
  • Calcium regulation
  • Epididymis
  • Epididymosomes
  • PMCA
  • Plasma membrane calcium pump
  • Sperm maturation

ASJC Scopus subject areas

  • Reproductive Medicine
  • Cell Biology

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    Patel, R., Al-Dossary, A. A., Stabley, D. L., Barone, C., Galileo, D. S., Strehler, E. E., & Martin-DeLeon, P. A. (2013). Plasma membrane Ca2+-ATPase 4 in murine epididymis: Secretion of splice variants in the luminal fluid and a role in sperm maturation. Biology of Reproduction, 89(1), [6]. https://doi.org/10.1095/biolreprod.113.108712