4.6 Article

Development of a novel spatiotemporal depletion system for cellular cholesterol

期刊

JOURNAL OF LIPID RESEARCH
卷 63, 期 3, 页码 -

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ELSEVIER
DOI: 10.1016/j.jlr.2022.100178

关键词

cholesterol; site-specific depletion; cholesterol quantification; cholesterol signaling; plasma membrane; lysosomes; chemically induced dimerization

资金

  1. National Institutes of Health [T32HL007820]

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Cholesterol is essential for mammalian cell membranes, but its site-specific functions have been limited by methodological constraints. A new cholesterol depletion system has been developed, allowing for spatial and temporal manipulation of cellular cholesterol levels.
Cholesterol is an essential component of mammalian cell membranes whose subcellular concentration and function are tightly regulated by de novo biosynthesis, transport, and storage. Although recent reports have suggested diverse functions of cellular cholesterol in different subcellular membranes, systematic investigation of its site-specific roles has been hampered by the lack of a methodology for spatiotemporal manipulation of cellular cholesterol levels. Here, we report the development of a new cholesterol depletion system that allows for spatiotemporal manipulation of intracellular cholesterol levels. This system utilizes a genetically encoded cholesterol oxidase whose intrinsic membrane binding activity is engineered in such a way that its membrane targeting can be controlled in a spatiotemporally specific manner via chemically induced dimerization. In combination with in situ quantitative imaging of cholesterol and signaling activity measurements, this system allows for unambiguous determination of site-specific functions of cholesterol in different membranes, including the plasma membrane and the lysosomal membrane.

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