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PMID: 24312544 Published · epublish English Journal Article Research Support, Non-U.S. Gov't

Protein phosphatases decrease their activity during capacitation: a new requirement for this event.

PloS one ·Vol. 8 ·No. 12 ·2013-00-00 ·Pages e81286

Signorelli JR, Díaz ES, Fara K, Barón L, Morales P

Abstract

There are few reports on the role of protein phosphatases during capacitation. Here, we report on the role of PP2B, PP1, and PP2A during human sperm capacitation. Motile sperm were resuspended in non-capacitating medium (NCM, Tyrode's medium, albumin- and bicarbonate-free) or in reconstituted medium (RCM, NCM plus 2.6% albumin/25 mM bicarbonate). The presence of the phosphatases was evaluated by western blotting and the subcellular localization by indirect immunofluorescence. The function of these phosphatases was analyzed by incubating the sperm with specific inhibitors: okadaic acid, I2, endothall, and deltamethrin. Different aliquots were incubated in the following media: 1) NCM; 2) NCM plus inhibitors; 3) RCM; and 4) RCM plus inhibitors. The percent capacitated sperm and phosphatase activities were evaluated using the chlortetracycline assay and a phosphatase assay kit, respectively. The results confirm the presence of PP2B and PP1 in human sperm. We also report the presence of PP2A, specifically, the catalytic subunit and the regulatory subunits PR65 and B. PP2B and PP2A were present in the tail, neck, and postacrosomal region, and PP1 was present in the postacrosomal region, neck, middle, and principal piece of human sperm. Treatment with phosphatase inhibitors rapidly (≤1 min) increased the percent of sperm depicting the pattern B, reaching a maximum of ∼40% that was maintained throughout incubation; after 3 h, the percent of capacitated sperm was similar to that of the control. The enzymatic activity of the phosphatases decreased during capacitation without changes in their expression. The pattern of phosphorylation on threonine residues showed a sharp increase upon treatment with the inhibitors. In conclusion, human sperm express PP1, PP2B, and PP2A, and the activity of these phosphatases decreases during capacitation. This decline in phosphatase activities and the subsequent increase in threonine phosphorylation may be an important requirement for the success of sperm capacitation.

MeSH Terms
Enzyme Inhibitors/pharmacology Humans Male Phosphoprotein Phosphatases/antagonists & inhibitors,chemistry,metabolism Phosphorylation/drug effects Sperm Capacitation/drug effects Spermatozoa/enzymology,physiology Threonine/metabolism
Chemicals
Enzyme Inhibitors Threonine Phosphoprotein Phosphatases
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Signorelli Janetti R
Department of Biomedicine, Faculty of Health Sciences, University of Antofagasta, Antofagasta, Chile.
Díaz Emilce S
Fara Karla
Barón Lina
Morales Patricio
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Article Info
Journal
PloS one
Abbr.
PLoS One
ISSN
1932-6203
DOI
10.1371/journal.pone.0081286
Published
2013-00-00
Epub
2013-00-02
Pages
e81286
Language
English
Region
United States
NLM ID
101285081
PMCID
PMC3846847
Subset
IM
External Links
PubMed source DOI: 10.1371/journal.pone.0081286 PMC Full Text View journal details
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