INVESTIGADORES
GALIGNIANA Mario Daniel
artículos
Título:
The role of DnaJ-like proteins in glucocorticoid receptor.hsp90 heterocomplex assembly by the reconstituted hsp90.p60.hsp70 foldosome complex
Autor/es:
DITTMAR KD, BANACH M, GALIGNIANA MD, PRATT WB
Revista:
JOURNAL OF BIOLOGICAL CHEMISTRY
Editorial:
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
Referencias:
Año: 1998 vol. 273 p. 7358 - 7366
ISSN:
0021-9258
Resumen:
The glucocorticoid receptor (GR) is recovered from hormone-free cells in
a heterocomplex with the molecular chaperone hsp90, which is required
to produce the proper folding state for steroid binding. GR.hsp90
heterocomplexes are formed by a multiprotein system that appears to
exist in all eukaryotic cells. Recently, we have reconstituted a
receptor.hsp90 heterocomplex assembly system with purified rabbit hsp90
and hsp70 and bacterially expressed human p23 and p60. We have shown
that hsp90, p60, and hsp70 form an hsp90.p60. hsp70 complex that
converts the GR from a non-steroid binding to a steroid binding form
(Dittmar, K. D., and Pratt, W. B. (1997) J. Biol. Chem. 272,
13047-13054). The resulting GR.hsp90 heterocomplex rapidly disassembles
unless p23 is present to bind to the ATP-dependent conformation of hsp90
and stabilize its association with the receptor (Dittmar, K. D.,
Demady, D. R., Stancato, L. F., Krishna, P., and Pratt, W. B. (1997) J.
Biol. Chem. 272, 21213-21220). In the current work, we show that the
purified rabbit hsp70 utilized in prior studies is contaminated with a
small amount of the rabbit DnaJ homolog hsp40. Elimination of the hsp40
from the purified GR.hsp90 assembly system reduces assembly activity,
and the activity is restored by addition of the purified yeast DnaJ
homolog YDJ-1. hsp40 is a component of the hsp90.p60.hsp70 foldosome
complex isolated from reticulocyte lysate with antibody against p60.
Under conditions that promote binding of p23 to hsp90 (elevated
temperature, ATP, Nonidet P-40, molybdate), a five-membered (p23.
hsp90.p60.hsp70.hsp40) complex of chaperone proteins is formed in
reticulocyte lysate or from purified proteins. The hsp40-free, purified
assembly system has a modest level of assembly activity that is
maximally potentiated by YDJ-1 when it is present at about one-twentieth
the concentration of hsp70. Although hsp40 is not in the final GR.hsp90
heterocomplex isolated from L cell cytosol, it is in the GR.hsp90
heterocomplex assembled in reticulocyte lysate. We conclude that hsp40
is a component of the multiprotein hsp90-based chaperone system where it
potentiates GR.hsp90 heterocomplex assembly.