title: Cryopreservation of common carp (Cyprinus carpio L.) sperm induces protein phosphorylation in tyrosine and threonine residues.
abstract: The effect of cryopreservation on the protein phosphorylation/dephosphorylation pattern of common carp (Cyprinus carpio) sperm is described. Sperm was diluted in dimethyl sulfoxide (DMSO) and ethylene glycol (EG)-based extenders, followed by equilibration, freezing, and thawing. Proteins extracted from fresh and cryopreserved spermatozoa were separated on SDS-PAGE and two-dimensional gel electrophoresis, blotted on polyvinylidene difluoride membrane, and treated with anti-phosphotyrosine, anti-phosphothreonine, or anti-phosphoserine antibodies. For the subsequent protein identification we used matrix-associated laser desorption/ionization time-of-flight mass spectrometry. The results demonstrated that cryopreservation with either DMSO or EG extender significantly altered the phosphorylation state of sperm proteins on tyrosine or threonine residues. A dramatic decrease in tyrosine phosphorylation was detected in the cryopreservation procedures with DMSO extender. Endoplasmin, transketolase, and S-adenosylhomocysteine hydrolase were identified as proteins that play a key role in cellular stress responses and oxidation and/or reduction reactions. Results indicate that the phosphorylation and/or dephosphorylation modifications of sperm proteins that occur during cryopreservation could stimulate a series of biochemical effects interfering with spermatozoa function and leading to a loss of motility and fertilization ability. Our findings indicated that use of EG extender provided superior protein preservation during sperm storage.
authors.affiliation: Faculty of Fisheries and Protection of Waters, South Bohemian Research Center of Aquaculture and Biodiversity of Hydrocenoses, University of South Bohemia in Ceske Budejovice, Vodnany, Czech Republic. pingli06@yahoo.com
authors.forename: P
authors.initials: P
authors.lastname: Li
authors.forename: M
authors.initials: M
authors.lastname: Hulak
authors.forename: Z H
authors.initials: ZH
authors.lastname: Li
authors.forename: M
authors.initials: M
authors.lastname: Sulc
authors.forename: M
authors.initials: M
authors.lastname: Psenicka
authors.forename: M
authors.initials: M
authors.lastname: Rodina
authors.forename: D
authors.initials: D
authors.lastname: Gela
authors.forename: O
authors.initials: O
authors.lastname: Linhart
mesh_terms: D000818:Animals; D002347:Carps / Q000502:physiology; D015925:Cryopreservation / Q000662:veterinary; D003451:Cryoprotective Agents / Q000494:pharmacology; D008297:Male; D010766:Phosphorylation / Q000187:drug effects; D011494:Protein Kinases / Q000378:metabolism*; D012662:Semen Preservation / Q000379:methods* / Q000662:veterinary; D013081:Sperm Motility / Q000187:drug effects; D013094:Spermatozoa / Q000187:drug effects / Q000502:physiology; D013912:Threonine / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects
chemical_list: D003451:Cryoprotective Agents; D013912:Threonine; D014443:Tyrosine; D011494:Protein Kinases