Positive and Negative Regulation of Cell Fusion in Saccharomyces cerevisiae

Abstract

Cell fusion is ubiquitous in eukaryotic organisms. Saccharomyces cerevisiae cells

fuse during mating; opposite mating types polarize their growth, come into contact,

forming the zone of cell fusion (ZCF), degrade their cell walls, and fuse their plasma

membranes. Because improper cell wall degradation would lead to cell lysis, the timing

and location of fusion is tightly regulated. Both positive and negative regulation of cell

wall degradation take place during yeast mating.

The highly conserved Rho-GTPase, Cdc42p, functions in polarized growth during

pheromone response and prezygote formation. Cdc42p binds the heterodimer

amphiphysin, Fus2p/Rvs161p, facilitating cell wall degradation. Cdc42p is recruited to a

focus at the center of the ZCF, dependent on Fus2p and ZCF curvature. Mutants with an

abnormally curved ZCF do not localize Cdc42p, suggesting that flatness of the ZCF

allows Fus2p/Rvs161p to recruit Cdc42p. In shmoos, Cdc42p neither forms a focus nor

colocalizes with Fus2.

A hyperactive component of the cell wall integrity (CWI) pathway, Pkc1p

(PKC1*), causes a cell fusion defect, suggesting a negative regulatory role in cell fusion.

The cell wall is monitored by five transmembrane proteins, Wsc1-3p, Mid2p, and Mtl1p,

which detect cell wall damage and activate the CWI pathway. Deletion of MID2 uniquely

causes pheromone-induced death, due to unregulated cell wall degradation. The

Cdc42p/Fus2p fusion complex forms in mid2 shmoos and mutations in cell fusion genes

(FUS1, FUS2, RVS161, CDC42) suppress mid2, implying that death is due to premature

cell fusion in the absence of a partner. PKC1* decreased Cdc42p focus formation in

prezygotes and mild over-expression of PKC1* suppressed the defects of both mid2Δ andwsc1Δ. Paradoxically, PKC1* increased Cdc42p focus formation in shmoos, neither colocalizing with Fus2p nor leading to premature lysis. We conclude that Mid2p and

Pkc1p negatively regulate cell fusion by disrupting the localization of Fus2p and/or its

interaction with Cdc42p.

In the course of these studies, we found that WSC1 has a zygote/diploid specific

role differing from the role of MID2. Diploids die at a high rate without WSC1, due to

cell lysis during budding. These differences underscore the distinct functions of two

similar CWI pathway mechanosensors, MID2 and WSC1.