Fluorescence visualization of receptor-bound low density lipoprotein in human fibroblasts

Richard G W Anderson, Joseph I. Goldstein, Michael S. Brown

Research output: Contribution to journalArticle

30 Citations (Scopus)

Abstract

Previous electron microscopic studies have shown that receptors for plasma low density lipoprotein (LDL) in human fibroblasts are located predominantly in coated pits on the cell surface. In the current studies, LDL receptors were visualized at the light microscopic level by two methods: 1) indirect immuno-fluorescent staining of receptor-bound LDL, and 2) direct visualization of LDL particles that were reconstituted with a fluorescent cholesteryl ester derivative. By both techniques, LDL receptors were seen to be clustered in minute foci that were aligned in linear arrays on the cell surface. These arrays corresponded to linear arrays of coated pits that were visualized by immunofluorescent staining of the coat protein, clathrin. During the internalization of receptor-bound LDL, each fluorescent focus of LDL particles seemed to enter the cell as an individual unit without the formation of large patches. Immediately after the reconstituted fluorescent LDL entered the cell, the fluorescent dots became larger in size and fewer in number, suggesting rapid fusion of endocytic vesicles. After internalization of LDL, new unoccupied receptors rapidly appeared on the cell surface where they were again distributed in linear arrays. In fibroblasts from a patient with the LDL internalization defect (J.D.), LDL receptors fail to be incorporated in coated pits. By immuno-fluorescence, LDL that was bound to these mutant receptors produced fluorescent foci that tended to be smaller and more numerous than those of normal fibroblasts. Moreover, the fluorescent foci of LDL

Original languageEnglish (US)
Pages (from-to)17-39
Number of pages23
JournalJournal of Receptors and Signal Transduction
Volume1
Issue number1
DOIs
StatePublished - 1980

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LDL Receptors
Fibroblasts
LDL Lipoproteins
Visualization
Fluorescence
Staining and Labeling
Transport Vesicles
Clathrin
Cholesterol Esters
Capsid Proteins
Electrons
Light
Fusion reactions
Derivatives
Plasmas
Defects

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology
  • Pharmacology

Cite this

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abstract = "Previous electron microscopic studies have shown that receptors for plasma low density lipoprotein (LDL) in human fibroblasts are located predominantly in coated pits on the cell surface. In the current studies, LDL receptors were visualized at the light microscopic level by two methods: 1) indirect immuno-fluorescent staining of receptor-bound LDL, and 2) direct visualization of LDL particles that were reconstituted with a fluorescent cholesteryl ester derivative. By both techniques, LDL receptors were seen to be clustered in minute foci that were aligned in linear arrays on the cell surface. These arrays corresponded to linear arrays of coated pits that were visualized by immunofluorescent staining of the coat protein, clathrin. During the internalization of receptor-bound LDL, each fluorescent focus of LDL particles seemed to enter the cell as an individual unit without the formation of large patches. Immediately after the reconstituted fluorescent LDL entered the cell, the fluorescent dots became larger in size and fewer in number, suggesting rapid fusion of endocytic vesicles. After internalization of LDL, new unoccupied receptors rapidly appeared on the cell surface where they were again distributed in linear arrays. In fibroblasts from a patient with the LDL internalization defect (J.D.), LDL receptors fail to be incorporated in coated pits. By immuno-fluorescence, LDL that was bound to these mutant receptors produced fluorescent foci that tended to be smaller and more numerous than those of normal fibroblasts. Moreover, the fluorescent foci of LDL",
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