Human spermatozoa contain multiple targets for protein S-nitrosylation: An alternative mechanism of the modulation of sperm function by nitric oxide?

TY - JOUR

T1 - Human spermatozoa contain multiple targets for protein S-nitrosylation

T2 - An alternative mechanism of the modulation of sperm function by nitric oxide?

AU - Lefièvre, Linda

AU - Chen, Yongjian

AU - Conner, Sarah J.

AU - Scott, Joanna L.

AU - Publicover, Steve J.

AU - Ford, W. Christopher L.

AU - Barratt, Christopher L. R.

PY - 2007/9

Y1 - 2007/9

N2 - Nitric oxide (NO) enhances human sperm motility and capacitation associated with increased protein phosphorylation. NO activates soluble guanylyl cyclase, but can also modify protein function covalently via S-nitrosylation of cysteine. Remarkably, this mechanism remains unexplored in sperm although they depend on post-translational protein modification to achieve changes in function required for fertilisation. Our objective was to identify targets for S-nitrosylation in human sperm. Spermatozoa were incubated with NO donors and S-nitrosylated proteins were identified using the biotin switch assay and a proteomic approach using MS/MS. 240 S-nitrosylated proteins were detected in sperm incubated with S-nitroso-glutathione. Minimal levels were observed in glutathione or untreated samples. Proteins identified consistently based on multiple peptides included established targets for S-nitrosylation in other cells e.g. tubulin, GST and HSPs but also novel targets including A-kinase anchoring protein (AKAP) types 3 and 4, voltage-dependent anion-selective channel protein 3 and semenogelin 1 and 2. In situ localisation revealed S-nitrosylated targets on the postacrosomal region of the head and throughout the flagellum. Potential targets for S-nitrosylation in human sperm include physiologically significant proteins not previously reported in other cells. Their identification will provide novel insight into the mechanism of action of NO in spermatozoa.

AB - Nitric oxide (NO) enhances human sperm motility and capacitation associated with increased protein phosphorylation. NO activates soluble guanylyl cyclase, but can also modify protein function covalently via S-nitrosylation of cysteine. Remarkably, this mechanism remains unexplored in sperm although they depend on post-translational protein modification to achieve changes in function required for fertilisation. Our objective was to identify targets for S-nitrosylation in human sperm. Spermatozoa were incubated with NO donors and S-nitrosylated proteins were identified using the biotin switch assay and a proteomic approach using MS/MS. 240 S-nitrosylated proteins were detected in sperm incubated with S-nitroso-glutathione. Minimal levels were observed in glutathione or untreated samples. Proteins identified consistently based on multiple peptides included established targets for S-nitrosylation in other cells e.g. tubulin, GST and HSPs but also novel targets including A-kinase anchoring protein (AKAP) types 3 and 4, voltage-dependent anion-selective channel protein 3 and semenogelin 1 and 2. In situ localisation revealed S-nitrosylated targets on the postacrosomal region of the head and throughout the flagellum. Potential targets for S-nitrosylation in human sperm include physiologically significant proteins not previously reported in other cells. Their identification will provide novel insight into the mechanism of action of NO in spermatozoa.

KW - Adaptor Proteins, Signal Transducing/metabolism

KW - Cysteine/metabolism

KW - Humans

KW - Male

KW - Mitochondrial Membrane Transport Proteins

KW - Nitric Oxide/metabolism

KW - Nitric Oxide Donors/metabolism

KW - Nitric Oxide Synthase Type III/metabolism

KW - Nitroso Compounds/metabolism

KW - Proteome/chemistry

KW - Proteomics/methods

KW - S-Nitrosoglutathione/metabolism

KW - Seminal Vesicle Secretory Proteins/metabolism

KW - Spermatozoa/metabolism

KW - Tandem Mass Spectrometry

KW - Voltage-Dependent Anion Channels/metabolism

U2 - 10.1002/pmic.200700254

DO - 10.1002/pmic.200700254

M3 - Article

C2 - 17683036

SN - 1615-9853

VL - 7

SP - 3066

EP - 3084

JO - Proteomics

JF - Proteomics

IS - 17

ER -