Regulation of Fab1 phosphatidylinositol 3-phosphate 5-kinase pathway by Vac7 protein and Fig4, a polyphosphoinositide phosphatase family member.
The Saccharomyces cerevisiae FAB1 gene encodes the sole phosphatidylinositol 3-phosphate [PtdIns(3)P] 5-kinase responsible for synthesis of the polyphosphoinositide PtdIns(3,5)P(2). VAC7 encodes a 128-kDa transmembrane protein that localizes to vacuolar membranes. Both vac7 and fab1 null mutants have dramatically enlarged vacuoles and cannot grow at elevated temperatures. Additionally, vac7Delta mutants have nearly undetectable levels of PtdIns(3,5)P(2), suggesting that Vac7 functions to regulate Fab1 kinase activity. To test this hypothesis, we isolated a fab1 mutant allele that bypasses the requirement for Vac7 in PtdIns(3,5)P(2) production. Expression of this fab1 allele in vac7Delta mutant cells suppresses the temperature sensitivity, vacuolar morphology, and PtdIns(3,5)P(2) defects normally exhibited by vac7Delta mutants. We also identified a mutant allele of FIG4, whose gene product contains a Sac1 polyphosphoinositide phosphatase domain, which suppresses vac7Delta mutant phenotypes. Deletion of FIG4 in vac7Delta mutant cells suppresses the temperature sensitivity and vacuolar morphology defects, and dramatically restores PtdIns(3,5)P(2) levels. These results suggest that generation of PtdIns(3,5)P(2) by the Fab1 lipid kinase is regulated by Vac7, whereas turnover of PtdIns(3,5)P(2) is mediated in part by the Sac1 polyphosphoinositide phosphatase family member Fig4.
Gary
Jonathan D
JD
Department of Cellular and Molecular Medicine, University of California at San Diego, School of Medicine, La Jolla, CA 92093-0668, USA.
Sato
Trey K
TK
Stefan
Christopher J
CJ
Bonangelino
Cecilia J
CJ
Weisman
Lois S
LS
Emr
Scott D
SD
D000483:Alleles; D000595:Amino Acid Sequence; D002462:Cell Membrane / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000378:metabolism; D005420:Flavoproteins / Q000378:metabolism*; D005656:Fungal Proteins / Q000378:metabolism*; D017353:Gene Deletion; D015971:Gene Expression Regulation, Enzymologic; D008565:Membrane Proteins / Q000378:metabolism*; D008954:Models, Biological; D008957:Models, Genetic; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010744:Phosphoric Monoester Hydrolases; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics* / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013997:Time Factors
D005420:Flavoproteins; D005656:Fungal Proteins; D008565:Membrane Proteins; D029701:Saccharomyces cerevisiae Proteins; C452436:VAC7 protein, S cerevisiae; C095504:FAB1 protein, S cerevisiae; D017853:Phosphotransferases (Alcohol Group Acceptor); C110669:FIG4 protein, S cerevisiae; D010744:Phosphoric Monoester Hydrolases