Conformational activation of a basic helix-loop-helix protein (MyoD1) by the C-terminal region of murine HSP90 (HSP84).

Abstract

A murine cardiac Agtll expression library was screened with an amphipathic helix antibody, and a recombinant representing the C-terminal 194 residues of murine HSP90 (HSP84) was cloned. Both recombinant and native HSP9Os were then found to rapidly convert a basic helix-loop-helix protein (MyoDi) from an inactive to an active conformation, as assayed by sequence-specific DNA binding. The conversion process involves a transient interaction between HSP9O and MyoDl and does not result in the formation of a stable tertiary complex. Conversion does not require ATP and occurs stoichiometrically in a dose-dependent fashion.HSP90 is an abundant, ubiquitous, and highly conserved protein present in most eukaryotic cells. These results provide direct evidence that HSP90 can affect the conformational structure of a DNA-binding protein.Heat shock proteins (HSPs) are thought to aid cells coping with stress, and their expression is induced by heat, heavy metals, pH, and alcohol (reviewed in references 13 and 16). Recent studies have shown that some classes of HSP (HSP60 and HSP70) also function in membrane translocation and folding of proteins in cells that are not under stress (reviewed in reference 13). Synthesis of HSP90 increases in some cells three to fivefold under stress conditions, but it is also abundant in uninduced cells (22). Most of the information acquired so far about the function of HSP90 has been through association, and an activity for the protein has not been described in vitro. HSP90 interacts with several viral oncogene products that display tyrosine kinase activity (4,25,31,39), associates with tubulin and actin (28, 36), stimulates the activity of eukarytotic initiation factor 2 a subunit-specific protein kinase (33), and forms stable complexes with some members of the nuclear receptor superfamily (see below). Sequence analysis has shown that HSP90 (the generic name for this family used here) is highly conserved among species (summarized in reference 15): human (HSP90), mouse (HSP84), chicken (HSP90), Drosophila (HSP83), Trypanosoma (HSP85), and yeast (HSP83) homologs display a minimum of 65% amino acid conservation.Members of the nuclear receptor superfamily display DNA-binding activity and translocate from the cytoplasm to the nucleus after acquiring a ligand. The inactive cytoplasmic forms of the glucocorticoid (18 [and references within], 35), estrogen (7 [and references within]), progesterone (6), and aryl hydrocarbon (AH) (29) receptors exist as stable complexes containing HSP90, whereas the ligand-activated nuclear forms lack HSP90. HSP90 is thought to negatively regulate activation and translocation of the cytoplasmic forms of the receptors in the absence of ligand (1). Substantial evidence suggests that binding of HSP90 may be essential to the subsequent activity of these receptors and that HSP90 may be involved in their initial folding. Glucocorticoid receptor translated from a reticulocyte lysate, which * Corresponding author.contains HSP90, displayed high-affinity hormone-binding activity, whe...

Hsp90 Is Required for Pheromone Signaling in Yeast

“…Similarly, many mutant alleles of the Drosophila HSP90 homolog, HSP83, are embryonic lethals over a deficiency of the locus (van der Straten et al, 1997), whereas the Escherichia coli homolog of Hsp90, HtpG, appears to be dispensable (Bardwell and Craig, 1988). Hsp90 can act as a molecular chaperone in vitro to promote refolding of denatured proteins (Wiech et al, 1992;Yonehara et al, 1996; see also Shaknovich et al, 1992;Shue and Kohtz, 1994), to hold denatured proteins in a folding-competent state for other chaperones (Freeman and Morimoto, 1996;Yonehara et al, 1996) and to prevent protein unfolding and aggregation (Miyata and Yahara, 1992;Jakob et al, 1995aJakob et al, , 1995bYonehara et al, 1996).…”

Low concentration of GA activates a preconditioning response in HepG2 cells during oxidative stress—roles of Hsp90 and vimentin

“…Hsp90 is a dimer and binds to several cellular proteins including steroid receptors and protein kinases (Jakob and Buchner 1994;Kimmins and MacRae 2000;Soti et al 1998). Hsp90 promotes folding of proteins (without adenosine triphosphate (ATP); Shaknovich et al 1992;Wiech et al 1992) and prevents protein unfolding and aggregation (Miyata and Yahara 1992;Jakob et al 1995a, b) by binding early unfolding intermediates (Jakob et al 1995a) and preventing their aggregation. Hsp90 also plays a very important role in quality control: it is required for degradation of certain misfolded substrates (McClellan et al 2005).…”

p53-independent association between SV40 large T antigen and the major cytosolic heat shock protein, HSP90

“…HSP90 has been suggested to modulate the function and subcellular distribution of target proteins. Functional folding, cellular localization, and DNA-binding of nuclear proteins have been shown to be regulated by HSP90 (Shaknovich et al, 1992;Hendrick and Hartl, 1993;Jakob and Buchner, 1994;Miyata and Yahara, 1995;Zou et al, 1998;Holt et al, 1999). We report here evidence that SV40 LT is associated with HSP90 in vivo, and that this association can be reconstituted from puri®ed HSP90 and puri®ed unfolded LT in vitro.…”

“…HSP70 is reported to a ect the structure of LT, and to complement the localization and functional defects of a LT mutant (Jeoung et al, 1991). HSP90 is also reported to a ect the conformation of several substrate proteins (Miyata and Yahara, 1992;Shaknovich et al, 1992;Wiech et al, 1992). Thus, HSP90 and HSP70 may both play important roles in the folding and nuclear translocation of LT in vivo.…”

“…HSP90 is known to interact with a variety of cellular and viral proteins (Rutherford and Zuker, 1994). The most well-characterized cellular targets of HSP90 are transcription factors, such as steroid hormone receptors (Joab et al, 1984;Catelli et al, 1985;Sanchez et al, 1985), MyoD1 (Shaknovich et al, 1992), and heat shock factor (Zou et al, 1998). Likely the largest family of HSP90-targets consists of protein kinases, such as casein kinase II (Miyata and Yahara, 1992), Raf1 (Stancato et al, 1993), sevenless tyrosine kinase (Cutforth and Rubin, 1994), and Wee1 (Aligue et al, 1994).…”