Heat-inducible degron: a method for constructing temperature-sensitive mutants.
A temperature-sensitive (ts) mutant retains the function of a gene at a low (permissive) temperature but not at a high (nonpermissive) temperature. Arg-DHFR, a dihydrofolate reductase bearing an amino-terminal (N-terminal) arginine, is long-lived in the yeast Saccharomyces cerevisiae, even though arginine is a destabilizing residue in the N-end rule of protein degradation. A ts derivative of Arg-DHFR was identified that is long-lived at 23 degrees C but rapidly degraded by the N-end rule pathway at 37 degrees C. Fusions of ts Arg-DHFR to either Ura3 or Cdc28 of S. cerevisiae confer ts phenotypes specific for these gene products. Thus, Arg-DHFRts is a heat-inducible degradation signal that can be used to produce ts mutants without a search for ts mutations.
Dohmen
R J
RJ
Division of Biology, California Institute of Technology, Pasadena 91125.
Wu
P
P
Varshavsky
A
A
D017489:CDC28 Protein Kinase, S cerevisiae / Q000235:genetics / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D006207:Half-Life; D006358:Hot Temperature; D016296:Mutagenesis; D010641:Phenotype; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics; D013696:Temperature; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics* / Q000378:metabolism; D014452:Ubiquitins
D005656:Fungal Proteins; D011993:Recombinant Fusion Proteins; D014452:Ubiquitins; D013762:Tetrahydrofolate Dehydrogenase; D017489:CDC28 Protein Kinase, S cerevisiae