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Membrane ruffling and macropinocytosis in A431 cells require cholesterol
Stine Grimmer1, Bo van Deurs, Kirsten Sandvig
1Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, 0310 Oslo, Norway.
Journal of Cell Science
|June 26, 2002
Summary
Cholesterol is essential for macropinosome formation. Depleting cholesterol prevents Rac1 protein localization to the cell membrane, inhibiting actin reorganization, membrane ruffling, and macropinocytosis.
Area of Science:
- Cell Biology
- Membrane Biology
- Biochemistry
Background:
- Cholesterol plays a role in forming caveolae and clathrin-coated pits.
- The role of cholesterol in macropinosome formation was previously unclear.
Purpose of the Study:
- To investigate the dependence of macropinosome formation on cholesterol in the plasma membrane.
- To elucidate the mechanism by which cholesterol influences macropinocytosis.
Main Methods:
- Macropinocytosis was induced in A431 cells using 12-O-tetradecanoylphorbol 13-acetate (PKC activator).
- Cholesterol was extracted using methyl-beta-cyclodextrin.
- Endocytosis was assessed by ricin uptake.
- Protein kinase C (PKC) activation and Rac1 localization were analyzed.
- Electron microscopy and confocal microscopy were employed.
Main Results:
- Cholesterol depletion inhibited phorbol ester-induced endocytosis of ricin.
- Extraction of cholesterol prevented membrane ruffles and macropinosome formation.
- Cholesterol depletion did not inhibit Rac1 activation but prevented its plasma membrane localization.
- The phorbol ester-induced reorganization of filamentous actin was inhibited by cholesterol depletion.
Conclusions:
- Cholesterol is required for the plasma membrane localization of activated Rac1.
- Cholesterol is essential for actin reorganization, membrane ruffling, and macropinocytosis.
- These findings highlight cholesterol's critical role in regulating endocytic processes.