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Mol. Hum. Reprod. Advance Access originally published online on March 21, 2007
Molecular Human Reproduction 2007 13(5):307-316; doi:10.1093/molehr/gam012
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© The Author 2007. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oxfordjournals.org
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Identification and localisation of SERCA 2 isoforms in mammalian sperm

C. Lawson1, V. Dorval1, S. Goupil1 and Pierre Leclerc1,2

1 Département d'Obstétrique/Gynécologie, Centre de Recherche en Biologie de la Reproduction, Université Laval and Ontogénie et Reproduction, Centre de recherche du CHUQ-CHUL, 2705 boul. Laurier, Sainte-Foy, Québec, QC, Canada G1V 4G2

2 To whom correspondence should be addressed at: Tel: +1-418-525-4444 ext. 46267; Fax: +1-418-654-2765; E-mail: pierre.leclerc{at}crchul.ulaval.ca

Upon binding to the egg's zona pellucida, capacitated spermatozoa will undergo a calcium-dependent exocytotic event called acrosome reaction. During this process, Ca2+ depletion from internal stores is followed by an important rise in [Ca2+]i due to a massive Ca2+ influx. Previous reports have shown that the acrosome can act as a Ca2+ store and that depletion of thapsigargin-sensitive stores induces acrosome exocytosis in capacitated spermatozoa from different mammalian species. The effect of thapsigargin, a specific inhibitor of sarcoplasmic/endoplasmic reticulum Ca2+-ATPases (SERCAs), suggests the presence and implication of SERCA in the active Ca2+ uptake during mammalian sperm capacitation. Although the presence of a thapsigargin-sensitive Ca2+-ATPase has been debated, the aim of this study was to clearly determine whether SERCAs are present in mammalian spermatozoa. Using three different anti-SERCA 2 antibodies, mono- and polyclonal, which recognised the same protein, we successfully identified and localised SERCA 2 in human, mouse and bovine sperm. Western blot analysis suggests that more than one SERCA 2 splice variant are present, one detected in the fraction containing the outer acrosomal membranes and another one present in the subcellular fraction containing the sperm midpiece. These results were confirmed by indirect immunofluorescence where SERCA 2 was observed in the acrosome and midpiece regions of human sperm. SERCA 2 immunohistochemical studies on human testis and PCR-amplification of mRNA encoding for each SERCA 2 splice variant in spermatogenic cells support the presence of this Ca2+-ATPase family in mature spermatozoa. In this paper, we clearly demonstrate, for the first time, the presence of SERCA 2 in mammalian sperm.

Key words: Ca2+-ATPase/calcium/capacitation/spermatozoa/intracellular stores

Submitted on December 8, 2006; resubmitted on January 30, 2007; accepted on February 2, 2007.


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