The nucleotide exchange factor Cdc24p may be regulated by auto-inhibition

Abstract

Site-specific activation of the Rho-type GTPase Cdc42p by its guanine-nucleotide exchange factor (GEF) Cdc24p is critical for the establishment of cell polarity. Here we show that binding of Cdc24p to the small GTPase Rsr1p/Bud1p is required for its recruitment to the incipient bud site. Rsr1p/Bud1p binds to the CH-domain of Cdc24p, which is essential for its function in vivo. We have identified a cdc24-mutant allele, which is specifically defective for bud-site selection. Our results suggest that Cdc24p is auto-inhibited by an intramolecular interaction with its carboxy-terminal PB1-domain. Rsr1p/Bud1p appears to activate the GEF activity of Cdc24p in vivo, possibly by triggering a conformational change that dissociates the PB1-domain from its intramolecular binding site. Genetic experiments suggest that Bem1p functions as a positive regulator of Cdc24p by binding to the PB1-domain of Cdc24p, thereby preventing its re-binding to the intramolecular inhibitory site. Taken together, our results support a two-step molecular mechanism for the site-specific activation of Cdc24p, which involves Rsr1p/Bud1p and the adaptor protein Bem1p.

Yeast and fungal morphogenesis from an evolutionary perspective

“…Thus the Rsr1/Bud5/ Bud2 GTPase module biases Cdc42-driven polarization toward the cortical landmark (bud scar). Consistent with the concept of "weak regulatory linkages", Rsr1 is not essential for Cdc24 activation or cortical recruitment and may be partially redundant with Bem1 in these activities [73,88].…”

“…During budding, Cdc24 is also a target of regulation dictating the orientation of bud formation relative to the previous bud scar (Figure 1). Cdc24 is activated and recruited to the plasma membrane partly through interaction with the GTP-bound Ras type GTPase Rsr1 [81][82][83]. Involvement of GTPase cascades is a recurring theme in spatial organization.…”

Activated Ezrin Promotes Cell Migration through Recruitment of the GEF Dbl to Lipid Rafts and Preferential Downstream Activation of Cdc42

“…This indicates that even the VAV2/Tiam1-promoted increase in Rac1 activity is not sufficient to promote cell spreading highlighting the significance of proper subcellular localization (Arthur et al, 2004). In Saccharomyces cerevisiae, targeting of the Cdc42p-specific GEF Cdc24p to the incipient bud site is essential for the recruitment and activation of Cdc42p, and deletion of the Dbl-homology domain responsible for targeting Cdc24p to the budding site is lethal and unable to complement the growth defects of Cdc24⌬ cells (Ziman and Johnson, 1994;Toenjes et al, 1999;Shimada et al, 2004). This indicates an evolutionarily conserved mechanism for proper localized GTPase activation.…”

Morphogenesis and the Cell Cycle

“…Localized GTP-Rsr1p in the vicinity of a landmark protein could recruit [and perhaps activate (Shimada et al 2004)] the GEF Cdc24p from the cytoplasm, leading to local GTP loading of Cdc42p at the membrane. This would create a local patch of GTP-Cdc42p in a sea of GDP-Cdc42p at the plasma membrane.…”