The upstream regulator, Rsr1p, and downstream effectors, Gic1p and Gic2p, of the Cdc42p small GTPase coordinately regulate initiation of budding in Saccharomyces cerevisiae.
BACKGROUND
Cdc42p, a Rho family small GTPase, is essential for budding initiation in the yeast Saccharomyces cerevisiae. The homologous proteins Gic1p and Gic2p (Gic1/2p) are effectors of Cdc42p, but their precise functions remain unknown. Rsr1p/Bud1p is a Ras family small GTPase that controls the selection of the budding site. Previous observations suggested that Rsr1p-GTP recruits Cdc24p, a GDP/GTP exchange factor for Cdc42p, at the incipient bud site. However, this model only addresses how Rsr1p determines the budding site, because the rsr1 mutant normally initiates budding.


RESULTS
Here we show that a rsr1 gic1 gic2 mutant fails to initiate budding, resulting in unbudded, large, and multinucleated cells. Expression of a dominant active or dominant negative mutant of RSR1 also inhibited the growth of the gic1 gic2 mutant, suggesting that cycling of Rsr1p between the GTP- and GDP-bound forms is required for budding initiation in the gic1 gic2 mutant. Among the mutations in effectors of CDC42, only the gic1 gic2 mutation demonstrated a synthetic lethal interaction with rsr1. Increased gene dosage of CDC42 suppressed defects in budding initiation of rsr1 gic1 gic2 mutants containing additional mutations in other effectors of CDC42, including BNI1, CLA4 or STE20. The polarized localization of Bni1p-GFP (green fluorescent protein) and Cla4p-GFP was lost after depletion of Gic1p in the rsr1 gic2 mutant.


CONCLUSIONS
We propose that Gic1/2p may stabilize or maintain a complex consisting of Cdc42p-GTP and its effectors at the budding site, which are assembled by the action of the Rsr1p-Cdc24p system.
Kawasaki
Ryosuke
R
Division of Molecular Interaction, Institute for Genetic Medicine, Hokkaido University Graduate School of Medicine, N15 W7, Kita-ku, Sapporo, 060-0815, Japan.
Fujimura-Kamada
Konomi
K
Toi
Hirofumi
H
Kato
Hiroyuki
H
Tanaka
Kazuma
K
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002455:Cell Division / Q000502:physiology*; D020549:GTP Phosphohydrolase Activators / Q000378:metabolism; D020558:GTP Phosphohydrolases; D020690:GTPase-Activating Proteins; D005804:Genes, Lethal; D020662:Guanine Nucleotide Exchange Factors; D009154:Mutation; D012441:Saccharomyces cerevisiae / Q000502:physiology; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D020846:cdc42 GTP-Binding Protein, Saccharomyces cerevisiae / Q000378:metabolism*; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; C083102:BUD2 protein, S cerevisiae; C091391:BUD5 protein, S cerevisiae; D002352:Carrier Proteins; C109252:GIC1 protein, S cerevisiae; C109253:GIC2 protein, S cerevisiae; D020549:GTP Phosphohydrolase Activators; D020690:GTPase-Activating Proteins; D020662:Guanine Nucleotide Exchange Factors; D029701:Saccharomyces cerevisiae Proteins; D020558:GTP Phosphohydrolases; C084013:RSR1 protein, S cerevisiae; D020846:cdc42 GTP-Binding Protein, Saccharomyces cerevisiae; D020691:rab GTP-Binding Proteins