title: In vivo bipartite interaction between the Hsp40 Sis1 and Hsp70 in Saccharomyces cerevisiae.
abstract: The essential Hsp40, Sis1, is a J-protein cochaperone for the Ssa class of Hsp70's of Saccharomyces cerevisiae. Sis1 is required for the maintenance of the prion [RNQ(+)], as Sis1 lacking its 55-amino-acid glycine-rich region (G/F) does not maintain [RNQ(+)]. We report that overexpression of Sis1DeltaG/F in an otherwise wild-type strain had a negative effect on both cell growth and [RNQ(+)] maintenance, while overexpression of wild-type Sis1 did not. Overexpression of the related Hsp40 Ydj1 lacking its G/F region did not cause inhibition of growth, indicating that this dominant effect of Sis1DeltaG/F is not a characteristic shared by all Hsp40's. Analysis of small deletions within the SIS1 G/F region indicated that the observed dominant effects were caused by the absence of sequences known to be important for Sis1's unique cellular functions. These inhibitory effects of Sis1DeltaG/F were obviated by alterations in the N-terminal J-domain of Sis1 that affect interaction with Ssa's ATPase domain. In addition, a genetic screen designed to isolate additional mutations that relieved these inhibitory effects identified two residues in Sis1's carboxy-terminal domain. These alterations disrupted the interaction of Sis1 with the 10-kD carboxy-terminal regulatory domain of Ssa1, indicating that Sis1 has a bipartite interaction with Ssa in vivo.
authors.affiliation: Department of Biochemistry, University of Wisconsin, Madison, 53706, USA.
authors.forename: Rebecca
authors.initials: R
authors.lastname: Aron
authors.forename: Nelson
authors.initials: N
authors.lastname: Lopez
authors.forename: William
authors.initials: W
authors.lastname: Walter
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Craig
authors.forename: Jill
authors.initials: J
authors.lastname: Johnson
mesh_terms: D000251:Adenosine Triphosphatases / Q000502:physiology; D000595:Amino Acid Sequence; D001667:Binding, Competitive; D049109:Cell Proliferation; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D005656:Fungal Proteins / Q000737:chemistry; D017353:Gene Deletion; D015966:Gene Expression Regulation, Fungal; D015723:Gene Library; D005800:Genes, Fungal; D005821:Genetic Techniques; D049452:Green Fluorescent Proteins / Q000378:metabolism; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins / Q000502:physiology*; D006360:Heat-Shock Proteins / Q000502:physiology*; D015151:Immunoblotting; D008156:Luciferases / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011328:Prions / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000502:physiology*; D013379:Substrate Specificity; D013696:Temperature; D013997:Time Factors
chemical_list: D005656:Fungal Proteins; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D011328:Prions; C070320:SIS1 protein, S cerevisiae; C090869:SSA3 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D049452:Green Fluorescent Proteins; D008156:Luciferases; D000251:Adenosine Triphosphatases; C095973:SSA1 protein, S cerevisiae