title: The Hsp70 homologue Lhs1p is involved in a novel function of the yeast endoplasmic reticulum, refolding and stabilization of heat-denatured protein aggregates.
abstract: Heat stress is an obvious hazard, and mechanisms to recover from thermal damage, largely unknown as of yet, have evolved in all organisms. We have recently shown that a marker protein in the ER of Saccharomyces cerevisiae, denatured by exposure of cells to 50 degrees C after preconditioning at 37 degrees C, was reactivated by an ATP-dependent machinery, when the cells were returned to physiological temperature 24 degrees C. Here we show that refolding of the marker enzyme Hsp150Delta-beta-lactamase, inactivated and aggregated by the 50 degrees C treatment, required a novel ER-located homologue of the Hsp70 family, Lhs1p. In the absence of Lhs1p, Hsp150Delta-beta-lactamase failed to be solubilized and reactivated and was slowly degraded. Coimmunoprecipitation experiments suggested that Lhs1p was somehow associated with heat-denatured Hsp150Delta- beta-lactamase, whereas no association with native marker protein molecules could be detected. Similar findings were obtained for a natural glycoprotein of S. cerevisiae, pro-carboxypeptidase Y (pro-CPY). Lhs1p had no significant role in folding or secretion of newly synthesized Hsp150Delta-beta-lactamase or pro-CPY, suggesting that the machinery repairing heat-damaged proteins may have specific features as compared to chaperones assisting de novo folding. After preconditioning and 50 degrees C treatment, cells lacking Lhs1p remained capable of protein synthesis and secretion for several hours at 24 degrees C, but only 10% were able to form colonies, as compared to wild-type cells. We suggest that Lhs1p is involved in a novel function operating in the yeast ER, refolding and stabilization against proteolysis of heatdenatured protein. Lhs1p may be part of a fundamental heat-resistant survival machinery needed for recovery of yeast cells from severe heat stress.
authors.affiliation: Institute of Biotechnology, University of Helsinki, Helsinki, Finland.
authors.forename: N
authors.initials: N
authors.lastname: Saris
authors.forename: H
authors.initials: H
authors.lastname: Holkeri
authors.forename: R A
authors.initials: RA
authors.lastname: Craven
authors.forename: C J
authors.initials: CJ
authors.lastname: Stirling
authors.forename: M
authors.initials: M
authors.lastname: Makarow
mesh_terms: D004721:Endoplasmic Reticulum / Q000378:metabolism*; D005656:Fungal Proteins / Q000378:metabolism; D006023:Glycoproteins; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism*; D006360:Heat-Shock Proteins / Q000378:metabolism*; D006358:Hot Temperature; D011485:Protein Binding; D011489:Protein Denaturation; D017510:Protein Folding; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D012995:Solubility
chemical_list: D005656:Fungal Proteins; D006023:Glycoproteins; C074229:HSP150 protein, S cerevisiae; D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; C102115:LHS1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins