The Hsp70-Ydj1 molecular chaperone represses the activity of the heme activator protein Hap1 in the absence of heme.
In Saccharomyces cerevisiae, heme directly mediates the effects of oxygen on transcription through the heme activator protein Hap1. In the absence of heme, Hap1 is bound by at least four cellular proteins, including Hsp90 and Ydj1, forming a higher-order complex, termed HMC, and its activity is repressed. Here we purified the HMC and showed by mass spectrometry that two previously unidentified major components of the HMC are the Ssa-type Hsp70 molecular chaperone and Sro9 proteins. In vivo functional analysis, combined with biochemical analysis, strongly suggests that Ssa proteins are critical for Hap1 repression in the absence of heme. Ssa may repress the activities of both Hap1 DNA-binding and activation domains. The Ssa cochaperones Ydj1 and Sro9 appear to assist Ssa in Hap1 repression, and only Ydj1 residues 1 to 172 containing the J domain are required for Hap1 repression. Our results suggest that Ssa-Ydj1 and Sro9 act together to mediate Hap1 repression in the absence of heme and that molecular chaperones promote heme regulation of Hap1 by a mechanism distinct from the mechanism of steroid signaling.
Hon
T
T
Department of Biochemistry, NYU School of Medicine, New York, New York 10016, USA.
Lee
H C
HC
Hach
A
A
Johnson
J L
JL
Craig
E A
EA
Erdjument-Bromage
H
H
Tempst
P
P
Zhang
L
L
D015153:Blotting, Western; D004268:DNA-Binding Proteins; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D006360:Heat-Shock Proteins / Q000378:metabolism*; D006418:Heme / Q000378:metabolism*; D046911:Macromolecular Substances; D013058:Mass Spectrometry; D008840:Microfilament Proteins / Q000172:deficiency / Q000378:metabolism; D018832:Molecular Chaperones / Q000378:metabolism*; D009154:Mutation; D016601:RNA-Binding Proteins; D012097:Repressor Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D017384:Sequence Deletion; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C058452:HAP1 protein, S cerevisiae; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D046911:Macromolecular Substances; D008840:Microfilament Proteins; D018832:Molecular Chaperones; D016601:RNA-Binding Proteins; D012097:Repressor Proteins; C110018:SRO9 protein, S cerevisiae; C542288:SSA2 protein, S cerevisiae; C090869:SSA3 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D014157:Transcription Factors; C070319:YDJ1 protein, S cerevisiae; D006418:Heme