GEF mechanism revealed by the structure of SmgGDS-558 and farnesylated RhoA complex and its implication for a chaperone mechanism

Abstract

SmgGDS has dual functions in cells and regulates small GTPases as both a guanine nucleotide exchange factor (GEF) for the Rho family and a molecular chaperone for small GTPases possessing a C-terminal polybasic region followed by four C-terminal residues called the CaaX motif, which is posttranslationally prenylated at its cysteine residue. Our recent structural work revealed that SmgGDS folds into tandem copies of armadillo-repeat motifs (ARMs) that are not present in other GEFs. However, the precise mechanism of GEF activity and recognition mechanism for the prenylated CaaX motif remain unknown because SmgGDS does not have a typical GEF catalytic domain and lacks a pocket to accommodate a prenyl group. Here, we aimed to determine the crystal structure of the SmgGDS/farnesylated RhoA complex. We found that SmgGDS induces a significant conformational change in the switch I and II regions that opens up the nucleotide-binding site, with the prenyl group fitting into the cryptic pocket in the N-terminal ARMs. Taken together, our findings could advance the understanding of the role of SmgGDS and enable drug design strategies for targeting SmgGDS and small GTPases.

Splice switching an oncogenic ratio of SmgGDS isoforms as a strategy to diminish malignancy

“…2A). Crystallography studies recently confirmed these structural differences in ARM domains between the 2 isoforms (26,27).…”

“…This possibility is supported by our finding that lowering the 607:558 ratio by overexpressing SmgGDS-558 induces apoptosis. SmgGDS-558 can promote the cytosolic accumulation of prenylated small GTPases (46)(47)(48), which occurs by SmgGDS-558 shielding the prenyl group of the GTPase from the aqueous environment of the cytosol (27). Other chaperones of prenylated GTPases, including PDEδ (49,50), PRA1 (49,51), VPS35 (52), and RhoGDI (53), similarly solubilize prenylated GTPases by shielding the prenyl group of the GTPase.…”

“…Our study describes the development of a SmgGDS-targeted approach for inhibiting the prenylation of oncogenic small GTPases. The crystal structure of SmgGDS was recently determined in complex with a small GTPase (26,27), making it more feasible to develop small-molecule inhibitors that target the binding surface between SmgGDS isoforms and small GTPase substrates to disrupt their prenylation and activation. Our unique strategy of inhibiting prenylation by redirecting the splicing of SmgGDS to reduce the 607:558 ratio highlights the effectiveness of SSOs in diminishing malignant phenotypes, and demonstrates how targeting SmgGDS to regulate small GTPase prenylation is a valid anticancer approach that should be further pursued.…”

Structural underpinnings of Ric8A function as a G-protein α-subunit chaperone and guanine-nucleotide exchange factor

“…Additional binding experiments revealed that the selectivity of the Ric8A/Gα interaction is determined, in part, by a few class-specific residues of Gα, particularly those at positions corresponding to I340 and N343 of Gα t . Armadillo proteins that are structurally related to Ric8A, including SmgGDS β-catenin, and importin-α, commonly bind their partners at the concave surface 41–43 . However, the nature of the ligand-binding interactions of Ric8A differs from those of other ARM-repeat proteins (Supplementary Fig.…”

Combined Proteomic and Genetic Interaction Mapping Reveals New RAS Effector Pathways and Susceptibilities

“…RAP1GDS1 also binds other small GTPases including KRAS (37). RAP1GDS1 differs structurally from canonical RHO and RAS GEFs (38) and may also act as a chaperone for these small GTPases (39). The CDKO screens point to specific GIs among RAP1GDS1, RHOA and mutant KRAS not previously reported.…”