A fluorescent cholesterol analog, R-Chol, mimics the dynamics of free cholesterol in live cells

IF 3 3区生物学 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY

Archives of biochemistry and biophysics Pub Date : 2025-08-23 DOI:10.1016/j.abb.2025.110600

Hinako Mogi, Yumi Yamashita, Masumi Sano, Ryo Kadowaki, Takuya Honda, Takafumi Kohama, Hiroyuki Nakamura

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Abstract

Cholesterol is an essential component of the mammalian cell membrane. Elucidating the dynamics of intracellular cholesterol is extremely important for understanding the mechanisms underlying life phenomena and diseases. The fluorescent cholesterol analog, R-Chol, has a fluorescent group at the 3-OH position of free cholesterol, which is less toxic and helpful for studying free cholesterol trafficking in live cells. In the present study, we determined the details of R-Chol dynamics in live cells. R-Chol was taken up by the plasma membrane within 10 min and then incorporated into the cells by endocytosis. Free cholesterol in late endosomes/lysosomes is transported to the endoplasmic reticulum via NPC1 proteins. In Npc1-deficient CHO cells, R-Chol accumulates in late endosomes/lysosomes. R-Chol accumulation in Npc1-deficient cells was attenuated by the overexpression of Rab9, which regulates intracellular transport from late endosomes to the Golgi apparatus. These results indicate that R-Chol incorporated via endocytosis is transported to late endosomes/lysosomes and then to the Golgi apparatus and endoplasmic reticulum by NPC1-dependent and NPC1-independent vesicular trafficking.

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一种荧光胆固醇类似物R-Chol可以模拟活细胞中游离胆固醇的动态变化

胆固醇是哺乳动物细胞膜的重要组成部分。阐明细胞内胆固醇的动力学对理解生命现象和疾病的机制至关重要。荧光胆固醇类似物R-Chol在游离胆固醇的3-OH位置有一个荧光基团，毒性较小，有助于研究活细胞中游离胆固醇的转运。在本研究中，我们确定了活细胞中R-Chol动力学的细节。R-Chol在10分钟内被质膜吸收，并通过内吞作用进入细胞。晚期内体/溶酶体中的游离胆固醇通过NPC1蛋白转运到内质网。在npc1缺失的CHO细胞中，R-Chol在晚期内体/溶酶体中积累。npc1缺陷细胞中的R-Chol积累通过Rab9的过表达而减弱，Rab9调节从内体晚期到高尔基体的细胞内运输。这些结果表明，通过内吞作用纳入的R-Chol通过npc1依赖性和非npc1依赖性的囊泡转运转运到晚期核内体/溶酶体，然后转运到高尔基体和内质网。

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来源期刊

Archives of biochemistry and biophysics 生物-生化与分子生物学

CiteScore

7.40

自引率

0.00%

发文量

245

审稿时长

26 days

期刊介绍： Archives of Biochemistry and Biophysics publishes quality original articles and reviews in the developing areas of biochemistry and biophysics. Research Areas Include: • Enzyme and protein structure, function, regulation. Folding, turnover, and post-translational processing • Biological oxidations, free radical reactions, redox signaling, oxygenases, P450 reactions • Signal transduction, receptors, membrane transport, intracellular signals. Cellular and integrated metabolism.