title: Src Kinase Is the Connecting Player between Protein Kinase A (PKA) Activation and Hyperpolarization through SLO3 Potassium Channel Regulation in Mouse Sperm.
abstract: Plasma membrane hyperpolarization is crucial for mammalian sperm to acquire acrosomal responsiveness during capacitation. Among the signaling events leading to mammalian sperm capacitation, the immediate activation of protein kinase A plays a pivotal role, promoting the subsequent stimulation of protein tyrosine phosphorylation that associates with fertilizing capacity. We have shown previously that mice deficient in the tyrosine kinase cSrc are infertile and exhibit improper cauda epididymis development. It is therefore not clear whether lack of sperm functionality is due to problems in epididymal maturation or to the absence of cSrc in sperm. To further address this problem, we investigated the kinetics of cSrc activation using anti-Tyr(P)-416-cSrc antibodies that only recognize active cSrc. Our results provide evidence that cSrc is activated downstream of PKA and that inhibition of its activity blocks the capacitation-induced hyperpolarization of the sperm plasma membrane without blocking the increase in tyrosine phosphorylation that accompanies capacitation. In addition, we show that cSrc inhibition also blocks the agonist-induced acrosome reaction and that this inhibition is overcome by pharmacological hyperpolarization. Considering that capacitation-induced hyperpolarization is mediated by SLO3, we evaluated the action of cSrc inhibitors on the heterologously expressed SLO3 channel. Our results indicate that, similar to SLO1 K(+) channels, cSrc blockers significantly decreased SLO3-mediated currents. Together, these results are consistent with findings showing that hyperpolarization of the sperm plasma membrane is necessary and sufficient to prepare the sperm for the acrosome reaction and suggest that changes in sperm membrane potential are mediated by cSrc activation.
authors.affiliation: From the Instituto de Biología Molecular y Celular de Rosario, CONICET-UNR, and Laboratorio de Especialidades Reproductivas, Facultad de Ciencias Bioquímicas y Farmacéuticas, UNR, Rosario SF2000, Argentina.
authors.forename: Cintia
authors.initials: C
authors.lastname: Stival
authors.affiliation: the Instituto de Biología y Medicina Experimental, CONICET, Ciudad Autónoma de Buenos Aires C1428ADN, Argentina.
authors.forename: Florenza A
authors.initials: FA
authors.lastname: La Spina
authors.affiliation: From the Instituto de Biología Molecular y Celular de Rosario, CONICET-UNR, and Laboratorio de Especialidades Reproductivas, Facultad de Ciencias Bioquímicas y Farmacéuticas, UNR, Rosario SF2000, Argentina.
authors.forename: Carolina
authors.initials: C
authors.lastname: Baró Graf
authors.affiliation: the Department of Veterinary and Animal Science, Paige Labs, University of Massachusetts, Amherst, Massachusetts 01003.
authors.forename: Enid
authors.initials: E
authors.lastname: Arcelay
authors.affiliation: From the Instituto de Biología Molecular y Celular de Rosario, CONICET-UNR, and Laboratorio de Especialidades Reproductivas, Facultad de Ciencias Bioquímicas y Farmacéuticas, UNR, Rosario SF2000, Argentina.
authors.forename: Silvia E
authors.initials: SE
authors.lastname: Arranz
authors.affiliation: the Department of Anatomy and Neurobiology, Washington University School of Medicine, St. Louis Missouri 63110, and.
authors.forename: Juan J
authors.initials: JJ
authors.lastname: Ferreira
authors.affiliation: the Department of Anatomy and Neurobiology, Washington University School of Medicine, St. Louis Missouri 63110, and.
authors.forename: Sibylle
authors.initials: S
authors.lastname: Le Grand
authors.affiliation: the Department of Anatomy and Neurobiology, Washington University School of Medicine, St. Louis Missouri 63110, and.
authors.forename: Victor A
authors.initials: VA
authors.lastname: Dzikunu
authors.affiliation: the Department of Anatomy and Neurobiology, Washington University School of Medicine, St. Louis Missouri 63110, and.
authors.forename: Celia M
authors.initials: CM
authors.lastname: Santi
authors.affiliation: the Department of Veterinary and Animal Science, Paige Labs, University of Massachusetts, Amherst, Massachusetts 01003.
authors.forename: Pablo E
authors.initials: PE
authors.lastname: Visconti
authors.affiliation: the Instituto de Biología y Medicina Experimental, CONICET, Ciudad Autónoma de Buenos Aires C1428ADN, Argentina.
authors.forename: Mariano G
authors.initials: MG
authors.lastname: Buffone
authors.affiliation: From the Instituto de Biología Molecular y Celular de Rosario, CONICET-UNR, and Laboratorio de Especialidades Reproductivas, Facultad de Ciencias Bioquímicas y Farmacéuticas, UNR, Rosario SF2000, Argentina, krapf@ibr-conicet.gov.ar.
authors.forename: Dario
authors.initials: D
authors.lastname: Krapf
mesh_terms: D000177:Acrosome / Q000378:metabolism; D000818:Animals; D002462:Cell Membrane / Q000235:genetics; D016764:Cell Polarity / Q000235:genetics; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D051036:Large-Conductance Calcium-Activated Potassium Channels / Q000235:genetics* / Q000378:metabolism; D008297:Male; D008564:Membrane Potentials / Q000235:genetics*; D051379:Mice; D015398:Signal Transduction / Q000235:genetics; D013075:Sperm Capacitation / Q000235:genetics; D013094:Spermatozoa / Q000378:metabolism; D019061:src-Family Kinases / Q000235:genetics / Q000378:metabolism*
chemical_list: C110627:Kcnu1 protein, mouse; D051036:Large-Conductance Calcium-Activated Potassium Channels; D000242:Cyclic AMP; D019061:src-Family Kinases; D017868:Cyclic AMP-Dependent Protein Kinases
keywords: PKA; Src; acrosome reaction; membrane potential; phosphotyrosine signaling; potassium channel; sperm