Author(s): Ho J, Bretscher A
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Abstract Polarized growth in yeast requires cooperation between the polarized actin cytoskeleton and delivery of post-Golgi secretory vesicles. We have previously reported that loss of the major tropomyosin isoform, Tpm1p, results in cells sensitive to perturbations in cell polarity. To identify components that bridge these processes, we sought mutations with both a conditional defect in secretion and a partial defect in polarity. Thus, we set up a genetic screen for mutations that conferred a conditional growth defect, showed synthetic lethality with tpm1Delta, and simultaneously became denser at the restrictive temperature, a hallmark of secretion-defective cells. Of the 10 complementation groups recovered, the group with the largest number of independent isolates was functionally null alleles of RAS2. Consistent with this, ras2Delta and tpm1Delta are synthetically lethal at 35 degrees C. We show that ras2Delta confers temperature-sensitive growth and temperature-dependent depolarization of the actin cytoskeleton. Furthermore, we show that at elevated temperatures ras2Delta cells are partially defective in endocytosis and show a delocalization of two key polarity markers, Myo2p and Cdc42p. However, the conditional enhanced density phenotype of ras2Delta cells is not a defect in secretion. All the phenotypes of ras2Delta cells can be fully suppressed by expression of yeast RAS1 or RAS2 genes, human Ha-ras, or the double disruption of the stress response genes msn2Deltamsn4Delta. Although the best characterized pathway of Ras function in yeast involves activation of the cAMP-dependent protein kinase A pathway, activation of the protein kinase A pathway does not fully suppress the actin polarity defects, suggesting that there is an additional pathway from Ras2p to Msn2/4p. Thus, Ras2p regulates cytoskeletal polarity in yeast under conditions of mild temperature stress through the stress response pathway.
This article was published in Mol Biol Cell
and referenced in Fungal Genomics & Biology