IBYME   02675
INSTITUTO DE BIOLOGIA Y MEDICINA EXPERIMENTAL
Unidad Ejecutora - UE
artículos
Título:
Molecular Cloning and Characterization of Phospholipase C Zeta in Equine Sperm and Testis Reveals Species-Specific Differences in Expression of Catalytically Active Protein
Autor/es:
BEDFORD-GUAUS SJ; MCPARTLIN LA; XIE J; WESTMILLER SL; BUFFONE MG; ROBERSON MS
Revista:
BIOLOGY OF REPRODUCTION
Editorial:
SOC STUDY REPRODUCTION
Referencias:
Año: 2010
ISSN:
0006-3363
Resumen:
Oocyte activation at fertilization is brought about by the
testis-specific phospholipase C zeta (PLCZ), owing to its ability to
induce intracellular Ca(2+) [Ca(2+)](i) oscillations. While this is a
highly conserved mechanism amongst mammals, important species-specific
differences in PLCZ sequence, activity and expression have been
reported. Thus the objectives of this research were to clone and
characterize the [Ca(2+)](i)-releasing activity and expression of equine
PLCZ in sperm and testis. Molecular cloning of equine PLCZ yielded a
1914 bp sequence that translated into a protein of the appropriate size
(~73kDa), as detected with an anti-PLCZ specific antibody.
Microinjection of 1 µg/µl equine PLCZ cRNA supported [Ca(2+)](i)
oscillations in murine oocytes that were of a higher relative frequency
than those generated by an equivalent concentration of murine Plcz cRNA.
Immunofluorescence revealed expression of PLCZ over the acrosome,
equatorial segment and head-midpiece junction; unexpectedly, PLCZ also
localized to the principal piece of the flagellum, in all epididymal,
uncapacitated and capacitated sperm. Immunostaining over the acrosome
was abrogated after induction of acrosomal exocytosis. Moreover,
injection of either sperm heads or tails into mouse oocytes showed that
PLCZ in both fractions is catalytically active. Immunohistochemistry on
equine testis revealed expression as early as the round spermatid stage
and injection of these cells supported [Ca(2+)](i) oscillations in
oocytes. In summary, we report that equine PLCZ displays higher
intrinsic [Ca(2+)](i)-releasing activity than murine PLCZ, and that
catalytically active protein is expressed in round spermatids as well as
the sperm flagellum, emphasizing important species-specific
differences. Moreover, some of these results may suggest potential novel
roles for PLCZ in sperm physiology.