Phorbol ester promotes endocytosis by activating a factor involved in endosome fusion.
Previous studies indicate that a zinc- and phorbol ester-binding factor is necessary for in vitro endosome fusion and for the effect of Rab5 on endosome fusion. Rab5 is a small GTPase that regulates membrane fusion between early endosomes derived from either receptor-mediated endocytosis or fluid-phase endocytosis. In its GTP-bound form, Rab5 promotes endocytosis and enhances fusion among early endosomes. To determine if PMA stimulates endocytosis by activating a factor required for endosome fusion, we overexpressed wild-type Rab5, a dominant negative mutant (Rab5:S34N), and a GTPase deficient mutant (Rab5:Q79L) in BHK-21 cells. The phorbol ester PMA stimulates endocytosis and increases the number and the size of endocytic vesicles, even in the presence of Rab5:S34N. Zinc depletion with N,N,N',N'-tetrakis-(2-pyridylmethyl)ethylenediamine (TPEN) and addition of calphostin C (CPC), an inhibitor of PKC that interacts with zinc and phorbol ester binding motifs, inhibited both basal and Rab5-stimulated fluid phase endocytosis. These two reagents also inhibited the size and number of endocytic vesicles promoted by Rab5. These results suggest that PMA stimulates endocytosis by regulating the dynamics of the early endosome compartment.
Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, MO 63110, USA.
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Aballay
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Stahl
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Mayorga
D000470:Alkaloids; D000818:Animals; D053119:Benzophenanthridines; D016466:CHO Cells; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D002614:Chelating Agents / Q000494:pharmacology; D006224:Cricetinae; D004705:Endocytosis / Q000187:drug effects / Q000502:physiology*; D011992:Endosomes / Q000187:drug effects / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005029:Ethylenediamines / Q000494:pharmacology; D020558:GTP Phosphohydrolases / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D008561:Membrane Fusion / Q000187:drug effects / Q000502:physiology; D016297:Mutagenesis, Site-Directed; D009281:Naphthalenes / Q000494:pharmacology; D010617:Phenanthridines / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D011994:Recombinant Proteins / Q000378:metabolism; D012845:Sindbis Virus / Q000235:genetics; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014162:Transfection; D015032:Zinc / Q000502:physiology; D020696:rab5 GTP-Binding Proteins
D000470:Alkaloids; D053119:Benzophenanthridines; D002614:Chelating Agents; D004791:Enzyme Inhibitors; D005029:Ethylenediamines; D009281:Naphthalenes; D010617:Phenanthridines; D011994:Recombinant Proteins; C016299:chelerythrine; D011493:Protein Kinase C; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; D020696:rab5 GTP-Binding Proteins; C058819:calphostin C; D015032:Zinc; D013755:Tetradecanoylphorbol Acetate; C044387:N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine