Mutational analysis of the beta-subunit of yeast geranylgeranyl transferase I.
The gene CAL1 (also known as CDC43) of Saccharomyces cerevisiae encodes the beta subunit of geranylgeranyl transferase I (GGTase I), which modifies several small GTPases. Biochemical analyses of the mutant enzymes encoded by cal1-1, and cdc43-2 to cdc43-7, expressed in bacteria, have shown that all of the mutant enzymes possess reduced activity, and that none shows temperature-sensitive enzymatic activities. Nonetheless, all of the cal1/cdc43 mutants show temperature-sensitive growth phenotypes. Increase in soluble pools of the small GTPases was observed in the yeast mutant cells at the restrictive temperature in vivo, suggesting that the yeast prenylation pathway itself is temperature sensitive. The cal1-1 mutation, located most proximal to the C-terminus of the protein, differs from the other cdc43 mutations in several respects. An increase in soluble Rho1p was observed in the cal1-1 strain grown at the restrictive temperature. The temperature-sensitive phenotype of cal1-1 is most efficiently suppressed by overproduction of Rho1p. Overproduction of the other essential target, Cdc42p, in contrast, is deleterious in cal1-1 cells, but not in other cdc43 mutants or the wild-type strains. The cdc43-5 mutant cells accumulate Cdc42p in soluble pools and cdc43-5 is suppressed by overproduction of Cdc42p. Thus, several phenotypic differences are observed among the cal1/cdc43 mutations, possibly due to alterations in substrate specificity caused by the mutations.
Ohya
Y
Y
Department of Biological Sciences, Graduate School of Science, University of Tokyo, Japan.
Caplin
B E
BE
Qadota
H
H
Tibbetts
M F
MF
Anraku
Y
Y
Pringle
J R
JR
Marshall
M S
MS
D019883:Alkyl and Aryl Transferases; D000595:Amino Acid Sequence; D001483:Base Sequence; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D004252:DNA Mutational Analysis; D004926:Escherichia coli / Q000235:genetics; D020558:GTP Phosphohydrolases / Q000235:genetics / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D005800:Genes, Fungal; D008969:Molecular Sequence Data; D009154:Mutation; D010641:Phenotype; D017368:Protein Prenylation; D011994:Recombinant Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D016415:Sequence Alignment; D013379:Substrate Specificity; D013489:Suppression, Genetic; D013696:Temperature; D014166:Transferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D020846:cdc42 GTP-Binding Protein, Saccharomyces cerevisiae; D020741:rho GTP-Binding Proteins
D018797:Cell Cycle Proteins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D014166:Transferases; D019883:Alkyl and Aryl Transferases; C083291:geranylgeranyltransferase type-I; C065203:CDC43 protein, S cerevisiae; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; C051302:RHO1 protein, S cerevisiae; D020846:cdc42 GTP-Binding Protein, Saccharomyces cerevisiae; D020741:rho GTP-Binding Proteins