Gef1p, a New Guanine Nucleotide Exchange Factor for Cdc42p, Regulates Polarity inSchizosaccharomyces pombe

Abstract

Schizosaccharomyces pombe cdc42+regulates cell morphology and polarization of the actin cytoskeleton. Scd1p/Ral1p is the only described guanine nucleotide exchange factor (GEF) for Cdc42p in S. pombe. We have identified a new GEF, named Gef1p, specifically regulating Cdc42p. Gef1p binds to inactive Cdc42p but not to other Rho GTPases in two-hybrid assays. Overexpression of gef1+increases specifically the GTP-bound Cdc42p, and Gef1p is capable of stimulating guanine nucleotide exchange of Cdc42p in vitro. Overexpression ofgef1+causes changes in cell morphology similar to those caused by overexpression of the constitutively active cdc42G12V allele. Gef1p localizes to the septum. gef1+deletion is viable but causes a mild cell elongation and defects in bipolar growth and septum formation, suggesting a role for Gef1p in the control of cell polarity and cytokinesis. The double mutant gef1Δ scd1Δ is not viable, indicating that they share an essential function as Cdc42p activators. However, both deletion and overexpression of either gef1+orscd1+causes different morphological phenotypes, which suggest different functions. Genetic evidence revealed a link between Gef1p and the signaling pathway of Shk1/Orb2p and Orb6p. In contrast, no genetic interaction between Gef1p and Shk2p-Mkh1p pathway was observed.

A novel interplay between GEFs orchestrates Cdc42 activity during cell polarity and cytokinesis

“…However, previous reports have shown that while gef1Δ mutants are mainly monopolar, about 40% of interphase cells show bipolar growth ( Figure 5B) (Coll et al, 2003;Das et al, 2012;Das et al, 2015). If Gef1 is required for Scd1 localization to the new end, how does bipolar growth occur in some gef1Δ cells?…”

“…Thus, we asked whether a similar interaction occurs at sites of polarized growth. Gef1 promotes bipolar growth in fission yeast (Coll et al, 2003;Das et al, 2012). Cells lacking gef1 show increased monopolarity, with polarized growth only occurring at the old end.…”

“…In most eukaryotes, Cdc42 is regulated by numerous GEFs and GAPs, which complicates our understanding of GTPase regulation (Bos et al, 2007). In the fission yeast Schizosaccharomyces pombe, Cdc42 is activated by two GEFs, Gef1 and Scd1 (Chang et al, 1994;Coll et al, 2003). The presence of only two Cdc42 GEFs, and the well-documented process of cell polarization makes fission yeast an excellent model system to understand the mechanistic details of cell shape establishment.…”

“…Gef1 and Scd1 are partially redundant but exhibit unique phenotypes when deleted (Chang et al, 1994;Coll et al, 2003), indicating that they may regulate Cdc42 in different manners. Cells lacking scd1 appear depolarized (Chang et al, 1994), and Scd1 oscillates between the two cell ends, much like Cdc42 (Das et al, 2012).…”

“…Cells lacking scd1 appear depolarized (Chang et al, 1994), and Scd1 oscillates between the two cell ends, much like Cdc42 (Das et al, 2012). In contrast, gef1 mutants become narrower and grow in a monopolar, rather than a bipolar, manner (Coll et al, 2003). Furthermore, Cdc42 activity is reduced at the new end in gef1 mutants (Das et al, 2012).…”