Analysis of a Chinese hamster ovary cell mutant with defective mobilization of cholesterol from the plasma membrane to the endoplasmic reticulum

Natalie L. Jacobs, Biree Andemariam, Kathryn W. Underwood, Krishnanchali Panchalingam, David Sternberg, Margaret Kielian, Laura Liscum

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

The factors involved in shuttling cholesterol among cellular membranes have not been defined. Using amphotericin B selection, we previously isolated Chinese hamster ovary cell mutants expressing defects in intracellular cholesterol transport. Complementation analysis among seven mutants identified one cell line, mutant 3-6, with a unique defect. The present analysis revealed three key features of mutant 3-6. First, the movement of cholesterol both from the endoplasmic reticulum and through lysosomes to the plasma membrane was normal. However, when intact 3-6 cells were treated with sphingomyelinase, movement of plasma membrane cholesterol to acyl CoA: cholesterol acyltransferase in the endoplasmic reticulum was defective. Cellular cholesterol was mobilized to this enzyme upon activation by 25- hydroxycholesterol. Second, mutant 3-6 did not utilize endogenously synthesized sterol or low density lipoprotein-derived cholesterol for growth as effectively as parental Chinese hamster ovary cells. Finally, despite normal movement of cholesterol to the plasma membrane, mutant 3-6 was amphotericin B resistant. The plasma membrane cholesterol content was normal as assessed by cholesterol oxidase treatment and Semliki Forest virus fusion, which suggests that the 3-6 mutation alters the organization of cholesterol in the plasma membrane. Our characterization of this mutant cell line should facilitate the identification of gene(s) required for this transport pathway.

Original languageEnglish (US)
Pages (from-to)1973-1987
Number of pages15
JournalJournal of Lipid Research
Volume38
Issue number10
StatePublished - Oct 1997

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Cell membranes
Cricetulus
Endoplasmic Reticulum
Ovary
Cholesterol
Cells
Cell Membrane
Amphotericin B
Cholesterol Oxidase
Sterol O-Acyltransferase
Semliki forest virus
Sphingomyelin Phosphodiesterase
Cell Line
Enzyme Activation
Defects
Sterols
Lysosomes
LDL Cholesterol
Viruses
LDL Lipoproteins

Keywords

  • Low density lipoprotein
  • Somatic cell mutant
  • Sphingomyelinase

ASJC Scopus subject areas

  • Endocrinology

Cite this

Jacobs, N. L., Andemariam, B., Underwood, K. W., Panchalingam, K., Sternberg, D., Kielian, M., & Liscum, L. (1997). Analysis of a Chinese hamster ovary cell mutant with defective mobilization of cholesterol from the plasma membrane to the endoplasmic reticulum. Journal of Lipid Research, 38(10), 1973-1987.

Analysis of a Chinese hamster ovary cell mutant with defective mobilization of cholesterol from the plasma membrane to the endoplasmic reticulum. / Jacobs, Natalie L.; Andemariam, Biree; Underwood, Kathryn W.; Panchalingam, Krishnanchali; Sternberg, David; Kielian, Margaret; Liscum, Laura.

In: Journal of Lipid Research, Vol. 38, No. 10, 10.1997, p. 1973-1987.

Research output: Contribution to journalArticle

Jacobs, NL, Andemariam, B, Underwood, KW, Panchalingam, K, Sternberg, D, Kielian, M & Liscum, L 1997, 'Analysis of a Chinese hamster ovary cell mutant with defective mobilization of cholesterol from the plasma membrane to the endoplasmic reticulum', Journal of Lipid Research, vol. 38, no. 10, pp. 1973-1987.
Jacobs, Natalie L. ; Andemariam, Biree ; Underwood, Kathryn W. ; Panchalingam, Krishnanchali ; Sternberg, David ; Kielian, Margaret ; Liscum, Laura. / Analysis of a Chinese hamster ovary cell mutant with defective mobilization of cholesterol from the plasma membrane to the endoplasmic reticulum. In: Journal of Lipid Research. 1997 ; Vol. 38, No. 10. pp. 1973-1987.
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