Use of dansyl-cholestanol as a probe of cholesterol behavior in membranes of living cells

作者：Huang Huan; McIntosh Avery L; Atshaves Barbara P; Ohno Iwashita Yoshiko; Kier Ann B; Schroeder Friedhelm^*

来源：The Journal of Lipid Research, 2010, 51(5): 1157-1172.

DOI：10.1194/jlr.M003244

• plasma membrane
• fluorescence
• imaging
• fibroblasts
• FRET
• affinity chromatography

While plasma membrane cholesterol-rich micro-domains play a role in cholesterol trafficking, little is known about the appearance and dynamics of cholesterol through these domains in living cells. The fluorescent cholesterol analog 6-dansyl-cholestanol (DChol), its biochemical fractionation, and confocal imaging of L-cell fibroblasts contributed the following new insights: i) fluorescence properties of DChol were sensitive to microenvironment polarity and mobility; (ii) DChol taken up by L-cell fibroblasts was distributed similarly as cholesterol and preferentially into cholesterol-rich vs. -poor microdomains resolved by affinity chromatography of purified plasma membranes; iii) DChol reported similar polarity ( dielectric constant near 18) but higher mobility near phospholipid polar head group region for cholesterol in purified cholesterol-rich versus -poor microdomains; and iv) real-time confocal imaging, quantitative colocalization analysis, and fluorescence resonance energy transfer with cholesterol-rich and -poor microdomain markers confirmed that DChol preferentially localized in plasma membrane cholesterol-rich microdomains of living cells. Thus, DChol sensed a unique, relatively more mobile microenvironment for cholesterol in plasma membrane cholesterol-rich microdomains, consistent with the known, more rapid exchange dynamics of cholesterol from cholesterol-rich than -poor microdomains.-Huang, H., A. L. McIntosh, B. P. Atshaves, Y. Ohno-Iwashita, A. B. Kier, and F. Schroeder. Use of dansyl-cholestanol as a probe of cholesterol behavior in membranes of living cells. J. Lipid Res. 2010. 51: 1157-1172.

• 出版日期2010-5