Energy-dependent conversion of transformed cytosolic glucocorticoid receptors from soluble to particulate-bound form.
We have recently published that soluble cytosolic glucocorticoid receptors are converted to a particulate form when they are incubated at 37 degrees C in a tubulin-polymerizing buffer [Pratt, W. B., Sanchez, E. R., Bresnick, E. H., Meshinchi, S., Scherrer, L. C., Dalman, F. C., & Welsh, M. J. (1989) Cancer Res. (Suppl.) 49, 2222s-2229s]. In this work, we further define this phenomenon and demonstrate that the L-cell glucocorticoid receptors are binding to a protein particulate composed largely of cytoskeletal proteins. Incubation of L-cell cytosol with glutamate at 37 degrees C converts the glucorticoid receptor to a form that pellets when cytosol is centrifuged at 150000g. The particulate material formed in a temperature-dependent and glutamate-dependent manner contains a large amount of tubulin, actin, and vimentin, but it is not the product of a cold-labile, colchicine-sensitive polymerization process. Very few cytosolic proteins are present in this complex, but the glucocorticoid receptor is tightly bound to it. Binding of the receptor to the cytoskeletal complex occurs after receptor transformation and is at least partially energy-dependent. Examination of the behavior of beta-galactosidase receptor fusion proteins and the nti glucocorticoid receptor demonstrates that residues 445 to the COOH-terminus of the receptor (DNA-binding and hormone-binding domains) contain the features required for binding to the cytoskeletal complex. Although it is the transformed receptor that associates tightly with the complex, DNA-binding activity is not required for association with the cytoskeletal particulate.(ABSTRACT TRUNCATED AT 250 WORDS)
Department of Pharmacology, University of Michigan Medical School, Ann Arbor 48109.
L C
LC
Scherrer
W B
WB
Pratt
D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D002498:Centrifugation; D003598:Cytoskeletal Proteins / Q000737:chemistry / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D005971:Glutamates / Q000494:pharmacology; D018698:Glutamic Acid; D006360:Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D007163:Immunosorbent Techniques; D051379:Mice; D008970:Molecular Weight; D011965:Receptors, Glucocorticoid / Q000737:chemistry* / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012995:Solubility; D013696:Temperature; D013816:Thermodynamics; D001616:beta-Galactosidase
D003598:Cytoskeletal Proteins; D005971:Glutamates; D006360:Heat-Shock Proteins; D011965:Receptors, Glucocorticoid; D011993:Recombinant Fusion Proteins; D018698:Glutamic Acid; D004247:DNA; D001616:beta-Galactosidase