Genetic evidence for phospholipid-mediated regulation of the Rab GDP-dissociation inhibitor in fission yeast.
We have previously identified mutant alleles of genes encoding two Rab proteins, Ypt3 and Ryh1, through a genetic screen using the immunosuppressant drug FK506 in fission yeast. In the same screen, we isolated gdi1-i11, a mutant allele of the essential gdi1+ gene encoding Rab GDP-dissociation inhibitor. In gdi1-i11, a conserved Gly267 was substituted by Asp. The Gdi1G267D protein failed to extract Rabs from membrane and Rabs were depleted from the cytosolic fraction in the gdi1-i11 mutant cells. Consistently, the Gdi1G267D protein was found mostly in the membrane fraction, whereas wild-type Gdi1 was found in both the cytosolic and the membrane fraction. Notably, overexpression of spo20+, encoding a phosphatidylcholine/phosphatidylinositol transfer protein, rescued gdi1-i11 mutation, but not ypt3-i5 or ryh1-i6. The gdi1-i11 and spo20-KC104 mutations are synthetically lethal, and the wild-type Gdi1 failed to extract Rabs from the membrane in the spo20-KC104 mutant. The phosphatidylinositol-transfer activity of Spo20 is dispensable for the suppression of the gdi1-i11 mutation, suggesting that the phosphatidylcholine-transfer activity is important for the suppression. Furthermore, knockout of the pct1+ gene encoding a choline phosphate cytidyltransferase rescued the gdi1-i11 mutation. Together, our findings suggest that Spo20 modulates Gdi1 function via regulation of phospholipid metabolism of the membranes.
Ma
Yan
Y
Division of Molecular Pharmacology and Pharmacogenomics, Department of Genome Sciences, Kobe University Graduate School of Medicine, Hyogo, Japan.
Kuno
Takayoshi
T
Kita
Ayako
A
Nabata
Toshiya
T
Uno
Satoshi
S
Sugiura
Reiko
R
D000483:Alleles; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001216:Asparagine / Q000378:metabolism; D017353:Gene Deletion; D005800:Genes, Fungal; D005816:Genetic Complementation Test; D006056:Golgi Apparatus / Q000648:ultrastructure; D020730:Guanine Nucleotide Dissociation Inhibitors / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010743:Phospholipids / Q000378:metabolism*; D012568:Schizosaccharomyces / Q000235:genetics* / Q000502:physiology* / Q000648:ultrastructure; D017386:Sequence Homology, Amino Acid; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*
C086796:GDP dissociation inhibitor 1; D020730:Guanine Nucleotide Dissociation Inhibitors; D010743:Phospholipids; D001216:Asparagine; D020691:rab GTP-Binding Proteins