Plasma membrane aminoglycerolipid flippase function is required for signaling competence in the yeast mating pheromone response pathway.
The class 4 P-type ATPases ("flippases") maintain membrane asymmetry by translocating phosphatidylethanolamine and phosphatidylserine from the outer leaflet to the cytosolic leaflet of the plasma membrane. In Saccharomyces cerevisiae, five related gene products (Dnf1, Dnf2, Dnf3, Drs2, and Neo1) are implicated in flipping of phosphatidylethanolamine, phosphatidylserine, and phosphatidylcholine. In MAT A: cells responding to α-factor, we found that Dnf1, Dnf2, and Dnf3, as well as the flippase-activating protein kinase Fpk1, localize at the projection ("shmoo") tip where polarized growth is occurring and where Ste5 (the central scaffold protein of the pheromone-initiated MAPK cascade) is recruited. Although viable, a MAT A: dnf1∆ dnf2∆ dnf3∆ triple mutant exhibited a marked decrease in its ability to respond to α-factor, which we could attribute to pronounced reduction in Ste5 stability resulting from an elevated rate of its Cln2⋅Cdc28-initiated degradation. Similarly, a MAT A: dnf1∆ dnf3∆ drs2∆ triple mutant also displayed marked reduction in its ability to respond to α-factor, which we could attribute to inefficient recruitment of Ste5 to the plasma membrane due to severe mislocalization of the cellular phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate pools. Thus proper remodeling of plasma membrane aminoglycerolipids and phosphoinositides is necessary for efficient recruitment, stability, and function of the pheromone signaling apparatus.
Division of Biochemistry, Biophysics and Structural Biology, Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720-3202.
Elodie
E
Sartorel
Division of Biochemistry, Biophysics and Structural Biology, Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720-3202.
Evelyne
E
Barrey
Division of Biochemistry, Biophysics and Structural Biology, Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720-3202.
Rebecca K
RK
Lau
Division of Biochemistry, Biophysics and Structural Biology, Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720-3202 jthorner@berkeley.edu.
Jeremy
J
Thorner
D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism*; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000235:genetics; D002462:Cell Membrane / Q000737:chemistry*; D010675:Pheromones / Q000502:physiology*; D010716:Phosphatidylinositols / Q000378:metabolism; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction
D018528:ATP-Binding Cassette Transporters; D048868:Adaptor Proteins, Signal Transducing; C104014:DRS2 protein, S cerevisiae; D010675:Pheromones; D010716:Phosphatidylinositols; C080321:STE5 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D011494:Protein Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); C489795:MSS4 protein, S cerevisiae; C533049:Fpk1 protein, S cerevisiae; D000251:Adenosine Triphosphatases; C480602:Dnf2 protein, S cerevisiae; D000252:Calcium-Transporting ATPases; C480601:Dnf1 protein, S cerevisiae