Isolation and proteomic analysis of mouse sperm detergent-resistant membrane fractions

Evidence for dissociation of lipid rafts during capacitation

Susan B. Sleight, Patricia V. Miranda, Nia Washington Plaskett, Bernhard Maier, Jeff Lysiak, Heidi Scrable, John C. Herr, Pablo E. Visconti

Research output: Contribution to journalArticle

124 Citations (Scopus)

Abstract

Mammalian sperm acquire fertilization capacity after residing in the female tract during a process known as capacitation. The present study examined whether cholesterol efflux during capacitation alters the biophysical properties of the sperm plasma membrane by potentially reducing the extent of lipid raft domains as analyzed by the isolation of detergent-resistant membrane fractions using sucrose gradients. In addition, this work investigated whether dissociation of the detergent-resistant membrane fraction during capacitation alters resident sperm raft proteins. Mouse sperm proteins associated with such fractions were studied by silver staining, tandem mass spectrometry, and Western blot analysis. Caveolin 1 was identified in sperm lipid rafts in multimeric states, including a high-molecular-weight oligomer that is sensitive to degradation under reducing conditions at high pH. Capacitation resulted in reduction of the light buoyant-density, detergent-resistant membrane fraction and decreased the array of proteins isolated within this fraction, including loss of the high-molecular-weight caveolin 1 oligomers. Proteomic analysis of sperm proteins isolated in the light buoyant-density fraction identified several proteins, including hexokinase 1, testis serine proteases 1 and 2, TEX101, hyaluronidase (PH20, SPAM1), facilitated glucose transporter 3, lactate dehydrogenase A, carbonic anhydrase IV, IZUMO, pantophysin, basigin, and cysteine-rich inhibitory secretory protein 1. Capacitation also resulted in a significant reduction of sperm labeling by the fluorescent lipid-analog DilC16, indicating that capacitation alters the liquid-ordered domains in the sperm plasma membrane. The observations that capacitation alters the protein composition of the detergent-resistant membrane fractions is consistent with the hypothesis that cholesterol efflux during capacitation dissociates lipid raft constituents, initiating signaling events that lead to sperm capacitation.

Original languageEnglish (US)
Pages (from-to)721-729
Number of pages9
JournalBiology of Reproduction
Volume73
Issue number4
DOIs
StatePublished - Oct 2005
Externally publishedYes

Fingerprint

Detergents
Proteomics
Spermatozoa
Lipids
Membranes
Caveolin 1
Proteins
Carbonic Anhydrase IV
CD147 Antigens
Molecular Weight
Cholesterol
Cell Membrane
Sperm Capacitation
Light
Protein Array Analysis
Silver Staining
Hexokinase
Hyaluronoglucosaminidase
Facilitative Glucose Transport Proteins
Serine Proteases

Keywords

  • Fertilization
  • Signal transduction
  • Sperm
  • Sperm capacitation
  • Testis

ASJC Scopus subject areas

  • Cell Biology
  • Developmental Biology
  • Embryology

Cite this

Isolation and proteomic analysis of mouse sperm detergent-resistant membrane fractions : Evidence for dissociation of lipid rafts during capacitation. / Sleight, Susan B.; Miranda, Patricia V.; Plaskett, Nia Washington; Maier, Bernhard; Lysiak, Jeff; Scrable, Heidi; Herr, John C.; Visconti, Pablo E.

In: Biology of Reproduction, Vol. 73, No. 4, 10.2005, p. 721-729.

Research output: Contribution to journalArticle

Sleight, Susan B. ; Miranda, Patricia V. ; Plaskett, Nia Washington ; Maier, Bernhard ; Lysiak, Jeff ; Scrable, Heidi ; Herr, John C. ; Visconti, Pablo E. / Isolation and proteomic analysis of mouse sperm detergent-resistant membrane fractions : Evidence for dissociation of lipid rafts during capacitation. In: Biology of Reproduction. 2005 ; Vol. 73, No. 4. pp. 721-729.
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