title: Hsp90 cochaperone Aha1 is a negative regulator of the Saccharomyces MAL activator and acts early in the chaperone activation pathway.
abstract: Aha1 is a ubiquitous cochaperone of the Hsp90/Hsp70 chaperone machine. It binds the middle domain of Hsp90 and stimulates ATPase activity, suggesting a function late in the chaperone pathway. Saccharomyces Mal63 MAL activator is a DNA-binding transcription factor and Hsp90 client protein. This study utilizes several MAL activator mutants to investigate Aha1 function in vivo. Deletion of AHA1 enhances induced Mal63-dependent maltase activity levels 2-fold, whereas overproduction of Aha1 represses expression. Maltase expression in strains carrying constitutive and super-inducible mutant activators with alterations near the C terminus (particularly residues 433-463) is unaffected by either aha1Delta or Aha1 overproduction. However, another constitutive activator with alterations outside of this C-terminal region is sensitive to Aha1 regulation. Previously, we showed that in the absence of inducer, Mal63 forms a stable intermediate complex with Hsp70, Hsp90, and Sti1, whereas noninducible mutant activators bind only with Hsp70 in an apparent early complex. Here, we find that triple Myc-tagged Aha1/Myc3 copurifies with all noninducible Mal63 mutant activators tested. Aha1/Myc3 association with inducible Mal63 is observed only in a sti1Delta strain, in which Hsp90 binding and intermediate complex formation are defective. Constitutive and super-inducible mutant activators with C-terminal alterations do not bind Aha1 even in a sti1Delta strain. Mal63 binding to Hsp90 and Hsp70 is enhanced 3-fold by loss of Aha1. These results suggest an interaction between Aha1 and residues near the C terminus of Mal63 thereby regulating Hsp90 association. A novel mechanism for the negative regulation of the MAL activator by Aha1 cochaperone is proposed.
authors.affiliation: Biology Department, Queens College-City University of New York, Flushing, New York 11367, USA.
authors.forename: Fulai
authors.initials: F
authors.lastname: Ran
authors.forename: Nidhi
authors.initials: N
authors.lastname: Gadura
authors.forename: Corinne A
authors.initials: CA
authors.lastname: Michels
mesh_terms: D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D018833:Chaperonins / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology / Q000528:radiation effects; D015966:Gene Expression Regulation, Fungal / Q000502:physiology; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D018841:HSP90 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D006360:Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D000520:alpha-Glucosidases / Q000096:biosynthesis / Q000235:genetics
chemical_list: C475092:AHA1 protein, S cerevisiae; D018840:HSP70 Heat-Shock Proteins; D018841:HSP90 Heat-Shock Proteins; D006360:Heat-Shock Proteins; C120379:MAL63 protein, S cerevisiae; D046912:Multiprotein Complexes; C542289:STI1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D000520:alpha-Glucosidases; D000251:Adenosine Triphosphatases; D018833:Chaperonins