Article ; Online: Bicarbonate-dependent serine/threonine protein dephosphorylation in capacitating boar spermatozoa.
2010 Volume 31, Issue 4, Page(s) 393–405
Abstract: This study investigates the dynamics of serine/threonine (S/T) protein phosphorylation in sperm incubated under capacitating (C) conditions using the boar as a model system. For the first time, this approach has identified multiple dephosphorylation ... ...
| Abstract | This study investigates the dynamics of serine/threonine (S/T) protein phosphorylation in sperm incubated under capacitating (C) conditions using the boar as a model system. For the first time, this approach has identified multiple dephosphorylation events that occur in a bicarbonate-dependent fashion. Different phospho-(S/T) kinase substrate antibodies were used, and dephosphorylation of 5 S/T phosphoproteins was observed in C sperm compared with noncapacitated (N) cells. Specifically, dephosphorylation of 96-, 90-, 64-, and 55-kd proteins was detected by immunoblotting using 2 phospho-Akt substrate antibodies and a phosphoprotein kinase A substrate antibody. In addition, dephosphorylation of a 105-kd protein was detected using a phospho-ATM/ATR substrate antibody. In contrast, no dephosphorylation was observed using a phosphoprotein kinase C substrate antibody, and increased tyrosine phosphorylation of 32- and 20-kd proteins was detected in C compared with N sperm. Immunolocalization experiments revealed subtle changes in the pattern expression as well as a reduction of phosphorylation in C sperm. Whereas sperm incubated in N medium containing dibutyryl cAMP (dbcAMP) and 3-isobutyl-1-methylxanthine (IBMX) did not show protein dephosphorylation, incubation in C medium with dbcAMP/IBMX showed dephosphorylation as well as increased phosphorylation of other proteins (p68, p51, and p29). Finally, calyculin A, a phosphatase inhibitor, prevented dephosphorylation of p96, p90, p64, and p55 but not p105. Based on these data, we propose 2 pathways of protein dephosphorylation that are active during capacitation and independent of cAMP. Together, this provides direct evidence for more complex S/T phosphorylation dynamics than has been previously described for sperm undergoing capacitation. |
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| MeSH term(s) | 1-Methyl-3-isobutylxanthine ; Animals ; Bicarbonates/metabolism ; Bucladesine ; Fluorescent Antibody Technique ; Immunoblotting ; Male ; Oxazoles ; Phosphorylation ; Protein Phosphatase 1/antagonists & inhibitors ; Protein Phosphatase 2/antagonists & inhibitors ; Protein-Serine-Threonine Kinases/metabolism ; Protein-Tyrosine Kinases/metabolism ; Sperm Capacitation ; Spermatozoa/enzymology ; Swine | |||||
| Chemical Substances | Bicarbonates ; Oxazoles ; Bucladesine (63X7MBT2LQ) ; calyculin A (7D07U14TK3) ; Protein-Tyrosine Kinases (EC 2.7.10.1) ; Protein-Serine-Threonine Kinases (EC 2.7.11.1) ; Protein Phosphatase 1 (EC 3.1.3.16) ; Protein Phosphatase 2 (EC 3.1.3.16) ; 1-Methyl-3-isobutylxanthine (TBT296U68M) | |||||
| Language | English | |||||
| Publishing date | 2010-07 | |||||
| Publishing country | United States | |||||
| Document type | Journal Article ; Research Support, Non-U.S. Gov't | |||||
| ZDB-ID | 604624-1 | |||||
| ISSN | 1939-4640 ; 0196-3635 | |||||
| ISSN (online) | 1939-4640 | |||||
| ISSN | 0196-3635 | |||||
| DOI | 10.2164/jandrol.109.008383 | |||||
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| Database | MEDical Literature Analysis and Retrieval System OnLINE |
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