Male Decapacitation Factor SPINK3 Blocks Membrane Hyperpolarization and Calcium Entry in Mouse Sperm.

Lucia Zalazar,Andreina Cesari,Cintia Stival,Anabella R Nicolli,Gerardo A De Blas,Dario Krapf

doi:10.3389/fcell.2020.575126

Abstract

Mammalian sperm acquire ability to fertilize through a process called capacitation, occurring after ejaculation and regulated by both female molecules and male decapacitation factors. Bicarbonate and calcium present in the female reproductive tract trigger capacitation in sperm, leading to acrosomal responsiveness and hyperactivated motility. Male decapacitating factors present in the semen avert premature capacitation, until detached from the sperm surface. However, their mechanism of action remains elusive. Here we describe for the first time the molecular basis for the decapacitating action of the seminal protein SPINK3 in mouse sperm. When present in the capacitating medium, SPINK3 inhibited Src kinase, a modulator of the potassium channel responsible for plasma membrane hyperpolarization. Lack of hyperpolarization affected calcium channels activity, impairing the acquisition of acrosomal responsiveness and blocking hyperactivation. Interestingly, SPINK3 acted only on non-capacitated sperm, as it did not bind to capacitated cells. Binding selectivity allows its decapacitating action only in non-capacitated sperm, without affecting capacitated cells.

Highlights

After testicular ejaculation, mammalian sperm transit the female reproductive tract where they undergo a series of biochemical and physiological modifications in order to gain fertilization competence
We have previously shown that the recombinant protein SPINK3 reduced acrosomal responsiveness in mouse sperm when added at the beginning of capacitation (Zalazar et al, 2012)
These results suggest that SPINK3 inhibitory effect on acrosome reaction (AR) is the consequence of sperm not achieving the capacitated state and not being able to acquire acrosome responsiveness

Summary

Introduction

Mammalian sperm transit the female reproductive tract where they undergo a series of biochemical and physiological modifications in order to gain fertilization competence. Capacitation is a highly complex process in which influx of HCO3− and Ca2+ plays a key role, acting on the atypical adenylyl cyclase sAC (aka ADCY10) (Hess et al, 2005; Stival et al, 2016) The activity of this cyclase increases intracellular cAMP concentration which promotes the direct activation of the Ser/Thr kinase Protein Kinase A (PKA), orchestrating different signaling cascades downstream. Its activation is involved in: membrane potential (Em) hyperpolarization and acquisition of acrosomal responsiveness, flagellar hyperactivation, intracellular alkalization, and phosphorylation of sperm proteins in Tyr residues, among other known effects (Puga Molina et al, 2018). Even though these processes differ in their kinetics, all of them involve phosphorylation cascades initiated by PKA. We have recently shown that PKA activity is necessary for Src activation, which in turn leads to Em hyperpolarization and acrosomal responsiveness in mouse sperm (Stival et al, 2015)

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Conclusion