The Cdc42 Binding and Scaffolding Activities of the Fission Yeast Adaptor Protein Scd2

Abstract

The small GTP-binding protein Cdc42, the guanine nucleotide exchange factor Scd1, the p21-activated kinase Shk1, and the adaptor protein Scd2 are involved in the Cdc42-dependent signaling cascade in fission yeast. In the present study, we analyzed the Cdc42 binding and scaffolding activities of Scd2 by co-precipitation assays. We found that two SH3-containing regions, amino acid residues 1-87 (CB1 (Cdc42-binding region 1)) and 110 -266 (CB2), of Scd2 can bind to the GTP-bound form of Cdc42. CB2 is cryptic because of the intramolecular binding between the SH3 domain in CB2 (SH3(C)) and the PX domain and binds to Cdc42 only when the Scd2 PB1 domain binds to the PC motif-containing region (residues 760 -872) of Scd1. This CB2⅐Cdc42 association, which would stabilize the open configuration of Scd2, enables the SH3(C) domain to bind to the polyproline motif of Shk1. We also found that the GTP-bound form of Cdc42 binds to the CRIB motif of Shk1 more strongly than to Scd2. Thus, Scd2 functions as a scaffold to form a protein complex, and the GTP-bound Cdc42 might be transferred effectively from the upstream activator Scd1 to the downstream effector Shk1 via Scd2.

Elongation Factor Ts Directly Facilitates the Formation and Disassembly of the Escherichia coli Elongation Factor Tu·GTP·Aminoacyl-tRNA Ternary Complex

“…In yeast, the GEF Scd1 chaperones its activated G-protein, Cdc42, to its effector Shk1 via interactions with Scd2 (71). GEFs for the monomeric G-protein Ral, RalGDS and Rlf, interact with the GTP-bound state of upstream G-proteins Ras and Rap through Ras association domains (3,72).…”

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

“…Both models propose that active Cdc42 participates in the generation of the positive feedback. In fission yeast, a similar ternary complex with the GEF Scd1, scaffold Scd2, and kinase Pak1 has been reported (Endo et al, 2003). However, it is not clear how this complex mediates a positive feedback loop during polarized growth.…”

“…The Cdc42 ternary complex consists of the GEF Scd1, the scaffold protein Scd2, and the downstream effector Pak1 kinase (Endo et al, 2003). Observations in budding yeast suggest that the PAK kinase may mediate GEF recruitment (Kozubowski et al, 2008).…”

Pob1 Participates in the Cdc42 Regulation of Fission Yeast Actin Cytoskeleton

“…Indeed, the mutated Cdc42-879 protein was unable to interact with Pob1 by two-hybrid analysis ( Figure 5C). As a positive control, we used Scd2, a Cdc42 scaffold protein (Chang et al, 1994;Endo et al, 2003), that interacted with both wild-type Cdc42 and mutated Cdc42-879. These results suggest that Pob1 interaction with Cdc42 is important for its localization and/or stability.…”