Mobilisation of Ca2+ stores and flagellar regulation in human sperm by S-nitrosylation: a role for NO synthesised in the female reproductive tract

TY - JOUR

T1 - Mobilisation of Ca2+ stores and flagellar regulation in human sperm by S-nitrosylation

T2 - a role for NO synthesised in the female reproductive tract

AU - Machado-Oliveira, Gisela

AU - Lefievre, Linda

AU - Ford, Christopher

AU - Herrero, M. Belen

AU - Barratt, Christopher

AU - Connolly, Thomas J.

AU - Nash, Katherine

AU - Morales-Garcia, Aduen

AU - Kirkman-Brown, Jackson

AU - Publicover, Steve

PY - 2008

Y1 - 2008

N2 - Generation of NO by nitric oxide synthase (NOS) is implicated in gamete interaction and fertilisation. Exposure of human spermatozoa to NO donors caused mobilisation of stored Ca2+ by a mechanism that did not require activation of guanylate cyclase but was mimicked by S-nitroso-glutathione (GSNO; an S-nitrosylating agent). Application of dithiothreitol, to reduce protein -SNO groups, rapidly reversed the actions of NO and GSNO on [Ca2+](i). The effects of NO, GSNO and dithiothreitol on sperm protein S-nitrosylation, assessed using the biotin switch method, closely paralleled their actions on [Ca2+](i). Immunofluorescent staining revealed constitutive and inducible NOS in human oviduct and cumulus (the cellular layer investing the oocyte). 4,5-diaminofluorescein (DAF) staining demonstrated production of NO by these tissues. Incubation of human sperm with oviduct explants induced sperm protein S-nitrosylation resembling that induced by NO donors and GSNO. Progesterone (a product of cumulus cells) also mobilises stored Ca2+ in human sperm. Pre-treatment of sperm with NO greatly enhanced the effect of progesterone on [Ca2+](i), resulting in a prolonged increase in flagellar excursion. We conclude that NO regulates mobilisation of stored Ca2+ in human sperm by protein S-nitrosylation, that this action is synergistic with that of progesterone and that this synergism is potentially highly significant in gamete interactions leading to fertilisation.

AB - Generation of NO by nitric oxide synthase (NOS) is implicated in gamete interaction and fertilisation. Exposure of human spermatozoa to NO donors caused mobilisation of stored Ca2+ by a mechanism that did not require activation of guanylate cyclase but was mimicked by S-nitroso-glutathione (GSNO; an S-nitrosylating agent). Application of dithiothreitol, to reduce protein -SNO groups, rapidly reversed the actions of NO and GSNO on [Ca2+](i). The effects of NO, GSNO and dithiothreitol on sperm protein S-nitrosylation, assessed using the biotin switch method, closely paralleled their actions on [Ca2+](i). Immunofluorescent staining revealed constitutive and inducible NOS in human oviduct and cumulus (the cellular layer investing the oocyte). 4,5-diaminofluorescein (DAF) staining demonstrated production of NO by these tissues. Incubation of human sperm with oviduct explants induced sperm protein S-nitrosylation resembling that induced by NO donors and GSNO. Progesterone (a product of cumulus cells) also mobilises stored Ca2+ in human sperm. Pre-treatment of sperm with NO greatly enhanced the effect of progesterone on [Ca2+](i), resulting in a prolonged increase in flagellar excursion. We conclude that NO regulates mobilisation of stored Ca2+ in human sperm by protein S-nitrosylation, that this action is synergistic with that of progesterone and that this synergism is potentially highly significant in gamete interactions leading to fertilisation.

KW - Calcium

KW - Cumulus

KW - Motility

KW - Nitric oxide

KW - Oviduct

KW - Sperm

KW - Human

KW - Nitric oxide synthase

KW - Mitochondrial permeability transition

KW - Adenine nucleotide translocase

KW - Sensitive guanylyl cyclase

KW - Human spermatozoa

KW - Acrosome reaction

KW - In vitro

KW - Cumulus cells

KW - L-arginine

KW - Embryonic development

U2 - 10.1242/dev.024521

DO - 10.1242/dev.024521

M3 - Article

C2 - 18842814

SN - 0950-1991

VL - 135

SP - 3677

EP - 3686

JO - Development

JF - Development

IS - 22

ER -