Cloning and characterization of CAP, the S. cerevisiae gene encoding the 70 kd adenylyl cyclase-associated protein.
Adenylyl cyclase from S. cerevisiae contains at least two subunits, a 200 kd catalytic subunit and a subunit with an apparent molecular size of 70 kd, which we now call CAP (cyclase-associated protein). We cloned a cDNA encoding CAP by screening a yeast cDNA expression library in E. coli with antisera raised against the purified protein. The cDNA contained an open reading frame capable of encoding a 526 amino acid protein that is not homologous to any sequences in the current data bases. Adenylyl cyclase activity in membranes from cells that lacked CAP was not stimulated by RAS2 proteins in vitro. These results suggest that CAP is required for at least some aspects of the RAS-responsive signaling system. Mutants lacking CAP had four additional phenotypes that appear to be unrelated to effects of the RAS/adenylyl cyclase pathway: the inability to grow on rich medium (YPD), temperature sensitivity on minimal medium, sensitivity to nitrogen starvation, and a swollen cell morphology.
Field
J
J
Cold Spring Harbor Laboratory, New York 11724.
Vojtek
A
A
Ballester
R
R
Bolger
G
G
Colicelli
J
J
Ferguson
K
K
Gerst
J
J
Kataoka
T
T
Michaeli
T
T
Powers
S
S
D000262:Adenylyl Cyclases / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000235:genetics; D004789:Enzyme Activation; D005656:Fungal Proteins / Q000378:metabolism; D005800:Genes, Fungal; D005838:Genotype; D008969:Molecular Sequence Data; D008970:Molecular Weight; D015345:Oligonucleotide Probes; D010641:Phenotype; D010957:Plasmids; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D018631:ras Proteins
D004271:DNA, Fungal; D005656:Fungal Proteins; D015345:Oligonucleotide Probes; D029701:Saccharomyces cerevisiae Proteins; C489015:RAS2 protein, S cerevisiae; D018631:ras Proteins; D000262:Adenylyl Cyclases