Saccharomyces cerevisiae Rot1 is an essential molecular chaperone in the endoplasmic reticulum.
Molecular chaperones prevent aggregation of denatured proteins in vitro and are thought to support folding of diverse proteins in vivo. Chaperones may have some selectivity for their substrate proteins, but knowledge of particular in vivo substrates is still poor. We here show that yeast Rot1, an essential, type-I ER membrane protein functions as a chaperone. Recombinant Rot1 exhibited antiaggregation activity in vitro, which was partly impaired by a temperature-sensitive rot1-2 mutation. In vivo, the rot1-2 mutation caused accelerated degradation of five proteins in the secretory pathway via ER-associated degradation, resulting in a decrease in their cellular levels. Furthermore, we demonstrate a physical and probably transient interaction of Rot1 with four of these proteins. Collectively, these results indicate that Rot1 functions as a chaperone in vivo supporting the folding of those proteins. Their folding also requires BiP, and one of these proteins was simultaneously associated with both Rot1 and BiP, suggesting that they can cooperate to facilitate protein folding. The Rot1-dependent proteins include a soluble, type I and II, and polytopic membrane proteins, and they do not share structural similarities. In addition, their dependency on Rot1 appeared different. We therefore propose that Rot1 is a general chaperone with some substrate specificity.
Graduate School of Biological Sciences, Nara Institute of Science and Technology, Nara 630-0192, Japan.
Masato
M
Takeuchi
Yukio
Y
Kimata
Kenji
K
Kohno
D004721:Endoplasmic Reticulum / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D008565:Membrane Proteins / Q000378:metabolism*; D008954:Models, Biological; D018832:Molecular Chaperones / Q000378:metabolism*; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature
C081933:KRE6 protein, S cerevisiae; D008565:Membrane Proteins; D018832:Molecular Chaperones; C523667:ROT1 protein, S cerevisiae; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins