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Numerous studies have shown that cholesterol is a major component of lipid rafts. The purpose of this study was to establish the effects of cholesterol enrichment on the composition and function of lipid rafts in the mature mouse testis. Isolated lipid rafts were prepared from adult mouse testes by sucrose density gradient centrifugation. In the testes of male mice, the level of cholesterol was approximately 100 times higher in lipid rafts than in the non-raft fractions. Consistent with this, cholesterol-enriched lipid rafts were substantially depleted of a number of proteins, including β1-adrenoceptors, β1-integrin, caveolin-1, flotillin-1, signal transduction proteins (Gp43, G-proteins, Rap1) and translational proteins (eIF4E, eEF2). Importantly, the levels of spingomyelin phosphatase-2 and Sgpl1 mRNAs, which are highly localized in lipid rafts, were substantially decreased in the testes of cholesterol-depleted mice. Interestingly, in parallel with the functional impairment of cholesterol-enriched lipid rafts, the expression levels of mRNAs for the transcription factors GATA-4, TCF7L1 and MTF-1 were also decreased in the testes of cholesterol-depleted mice. Our data demonstrate that in the
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