Thundathil, JacobUnnikrishnan, Veena2022-11-152020-10-15Unnikrishnan, V. (2020). Ouabain-induced activation of Phospholipase C zeta and its contributions to bovine sperm capacitation (Master's thesis, University of Calgary, Calgary, Canada). Retrieved from https://prism.ucalgary.ca.http://hdl.handle.net/1880/115455https://dx.doi.org/10.11575/PRISM/40422The objective was to investigate involvement of testis-specific isoform of Na+/K+-ATPase ?4 (ATP1A4) in Phospholipase C zeta (PLC ?) activation during sperm capacitation and contributions of PLC ? activity to the capacitation process. Immunoprecipitation studies were done to confirm the interaction of PLC ? with ATP1A4 and to determine tyrosine phosphorylation of PLC ? during capacitation. Contributions of activated PLC ? to capacitation process were evaluated by determining phosphotyrosine content of sperm proteins (Immunoblotting), proportion of sperm acquiring hyperactivated motility (Computer-Assisted Sperm Analysis) and quantifying filamentous actin (F-actin; Flow cytometry analysis) content. Our results demonstrated that capacitation conditions promoted interaction of ATP1A4 with PLC ?, resulting in tyrosine phosphorylation of PLC ?. Incubation of sperm under capacitation conditions upregulated PLC-mediated biochemical modifications (hyperactive motility, tyrosine phosphoprotein content and increased F-actin content), suggesting that capacitation enhanced PLC activity. We inferred that interaction of ATP1A4 with PLC ? induces tyrosine phosphorylation of PLC ?, enhancing its activity and upregulation of capacitation-associated biochemical events. A capacitation-associated increase in PLC ? activity may enable sperm to initiate oocyte activation immediately after oocyte penetration (during fertilization). Since failure of oocyte activation after sperm penetration is considered a major reason for fertilization failure, results of this study may have important implications for male factor infertility treatments, e.g. improving efficacy of intracytoplasmic sperm injection (ICSI).enUniversity of Calgary graduate students retain copyright ownership and moral rights for their thesis. You may use this material in any way that is permitted by the Copyright Act or through licensing that has been assigned to the document. For uses that are not allowable under copyright legislation or licensing, you are required to seek permission.BovinecapaciatationfertilityOuabainPLC zetaBiological Sciencesmaster thesis