Disruption of the Raf-1-Hsp90 molecular complex results in destabilization of Raf-1 and loss of Raf-1-Ras association.
Cytosolic Raf-1 exists in a high molecular weight complex with the heat shock protein Hsp90, the purpose of which is unknown. The benzoquinone ansamycin, geldanamycin, specifically binds to Hsp90 and disrupts certain multimolecular complexes containing this protein. Using this drug, we are able to demonstrate rapid dissociation of both Raf-1-Hsp90 and Raf-1-Ras multimolecular complexes, concomitant with a markedly decreased half-life of the Raf-1 protein. Continued disruption of the Raf-1-Hsp90 complex results in apparent loss of Raf-1 protein from the cell, although Raf-1 synthesis is actually increased. Prevention of Raf-1-Hsp90 complex formation interferes with trafficking of newly synthesized Raf-1 from cytosol to plasma membrane. These data indicate that association with Hsp90 is essential for both Raf-1 protein stability and its proper localization in the cell.
Schulte
T W
TW
Clinical Pharmacology Branch, NCI, National Institutes of Health, Bethesda, Maryland 20892, USA.
Blagosklonny
M V
MV
Ingui
C
C
Neckers
L
L
D016475:3T3 Cells; D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D016227:Benzoquinones; D015153:Blotting, Western; D001943:Breast Neoplasms; D002460:Cell Line; D003600:Cytosol / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D018841:HSP90 Heat-Shock Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D047029:Lactams, Macrocyclic; D008715:Methionine / Q000378:metabolism; D051379:Mice; D008970:Molecular Weight; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D019908:Proto-Oncogene Proteins c-raf; D011809:Quinones / Q000494:pharmacology; D013462:Sulfur Radioisotopes; D013997:Time Factors; D014407:Tumor Cells, Cultured
D000903:Antibiotics, Antineoplastic; D016227:Benzoquinones; D018841:HSP90 Heat-Shock Proteins; D047029:Lactams, Macrocyclic; D011518:Proto-Oncogene Proteins; D011809:Quinones; D013462:Sulfur Radioisotopes; D008715:Methionine; D017346:Protein Serine-Threonine Kinases; D019908:Proto-Oncogene Proteins c-raf; C001277:geldanamycin