Expression and secretion of plasma membrane Ca2+-ATPase 4a (PMCA4a) during murine estrus

Association with oviductal exosomes and uptake in sperm

Amal A. Al-Dossary, Emanuel E. Strehler, Patricia A. Martin-DeLeon

Research output: Contribution to journalArticle

47 Citations (Scopus)

Abstract

PMCA4, a membrane protein, is the major Ca2+ efflux pump in murine sperm where its deletion leads to a severe loss of hyperactivated motility and to male infertility. We have previously shown that the PMCA4b splice variant interacts with CASK (Ca2+/CaM-dependent serine kinase) in regulating sperm Ca2+. More recently we detected that PMCA4a isoform, in addition to its presence in testis, is secreted in the epididymal luminal fluid and transferred to sperm. Here we show that Pmca4 mRNA is expressed in both the 4a and 4b variants in the vagina, uterus, and oviduct. Immunofluorescence reveals that PMCA4a is similarly expressed and is elevated during estrus, appearing in the glandular and luminal epithelia. Western analysis detected PMCA4a in all tissues and in the luminal fluids (LF) of the vagina (VLF), uterus (ULF), and the oviduct (OLF) collected during estrus. It was ∼9- and 4-fold higher in OLF than in VLF and ULF, and only marginally present in LF collected at metestrus/diestrus. Fractionation of the LF collected at estrus, via ultracentrifugation, revealed that 100% of the PMCA4a resides in the vesicular fraction of the ULF and OLF. Transmission electron microscopy (TEM) revealed that OLF vesicles have an exosomal orientation (with the cytoplasmic-side inward), a size range of 25-100 nm, with the characteristic CD9 biomarker. Thus, we dubbed these vesicles "oviductosomes", to which PMCA4a was immunolocalized. Incubation of caudal sperm in the combined LF or exosomes resulted in up to a ∼3-fold increase of sperm PMCA4a, as detected by flow cytometry, indicating in vitro uptake. Our results are consistent with the increased requirement of Ca2+ efflux in the oviduct. They show for the first time the presence of oviductal exosomes and highlight their role, along with uterosomes and vaginal exosomes, in post-testicular sperm acquisition of PMCA4a which is essential for hyperactivated motility and fertility. Copyright:

Original languageEnglish (US)
Article numbere80181
JournalPLoS One
Volume8
Issue number11
DOIs
StatePublished - Nov 14 2013

Fingerprint

exosomes
Exosomes
Ca2-transporting ATPase
Calcium-Transporting ATPases
Estrus
Cell membranes
Spermatozoa
estrus
plasma membrane
spermatozoa
Cell Membrane
secretion
uptake mechanisms
mice
Oviducts
oviducts
Fluids
calcium
vagina
Vagina

ASJC Scopus subject areas

  • Agricultural and Biological Sciences(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Medicine(all)

Cite this

Expression and secretion of plasma membrane Ca2+-ATPase 4a (PMCA4a) during murine estrus : Association with oviductal exosomes and uptake in sperm. / Al-Dossary, Amal A.; Strehler, Emanuel E.; Martin-DeLeon, Patricia A.

In: PLoS One, Vol. 8, No. 11, e80181, 14.11.2013.

Research output: Contribution to journalArticle

Al-Dossary, Amal A. ; Strehler, Emanuel E. ; Martin-DeLeon, Patricia A. / Expression and secretion of plasma membrane Ca2+-ATPase 4a (PMCA4a) during murine estrus : Association with oviductal exosomes and uptake in sperm. In: PLoS One. 2013 ; Vol. 8, No. 11.
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