title: The chaperone protein SmgGDS interacts with small GTPases entering the prenylation pathway by recognizing the last amino acid in the CAAX motif.
abstract: Ras family small GTPases localize at the plasma membrane, where they can activate oncogenic signaling pathways. Understanding the mechanisms that promote membrane localization of GTPases will aid development of new therapies to inhibit oncogenic signaling. We previously reported that SmgGDS splice variants promote prenylation and trafficking of GTPases containing a C-terminal polybasic region and demonstrated that SmgGDS-607 interacts with nonprenylated GTPases, whereas SmgGDS-558 interacts with prenylated GTPases in cells. The mechanism that SmgGDS-607 and SmgGDS-558 use to differentiate between prenylated and nonprenylated GTPases has not been characterized. Here, we provide evidence that SmgGDS-607 associates with GTPases through recognition of the last amino acid in the CAAX motif. We show that SmgGDS-607 forms more stable complexes in cells with nonprenylated GTPases that will become geranylgeranylated than with nonprenylated GTPases that will become farnesylated. These binding relationships similarly occur with nonprenylated SAAX mutants. Intriguingly, farnesyltransferase inhibitors increase the binding of WT K-Ras to SmgGDS-607, indicating that the pharmacological shunting of K-Ras into the geranylgeranylation pathway promotes K-Ras association with SmgGDS-607. Using recombinant proteins and prenylated peptides corresponding to the C-terminal sequences of K-Ras and Rap1B, we found that both SmgGDS-607 and SmgGDS-558 directly bind the GTPase C-terminal region, but the specificity of the SmgGDS splice variants for prenylated versus nonprenylated GTPases is diminished in vitro. Finally, we present structural homology models and data from functional prediction software to define both similar and unique features of SmgGDS-607 when compared with SmgGDS-558.
authors.affiliation: Department of Pharmacology and Toxicology, Medical College of Wisconsin, Milwaukee, Wisconsin 53226.
authors.forename: Nathan J
authors.initials: NJ
authors.lastname: Schuld
authors.affiliation: Department of Chemistry, University of Minnesota, Minneapolis, Minnesota 55414.
authors.forename: Jeffrey S
authors.initials: JS
authors.lastname: Vervacke
authors.affiliation: Department of Pharmacology and Toxicology, Medical College of Wisconsin, Milwaukee, Wisconsin 53226.
authors.forename: Ellen L
authors.initials: EL
authors.lastname: Lorimer
authors.affiliation: Department of Microbiology, Immunology, and Molecular Genetics, Medical College of Wisconsin, Milwaukee, Wisconsin 53226.
authors.forename: Nathan C
authors.initials: NC
authors.lastname: Simon
authors.affiliation: Department of Radiation Oncology, Medical College of Wisconsin, Milwaukee, Wisconsin 53226.
authors.forename: Andrew D
authors.initials: AD
authors.lastname: Hauser
authors.affiliation: Department of Microbiology, Immunology, and Molecular Genetics, Medical College of Wisconsin, Milwaukee, Wisconsin 53226.
authors.forename: Joseph T
authors.initials: JT
authors.lastname: Barbieri
authors.affiliation: Department of Chemistry, University of Minnesota, Minneapolis, Minnesota 55414.
authors.forename: Mark D
authors.initials: MD
authors.lastname: Distefano
authors.affiliation: Department of Pharmacology and Toxicology, Medical College of Wisconsin, Milwaukee, Wisconsin 53226. Electronic address: williams@mcw.edu.
authors.forename: Carol L
authors.initials: CL
authors.lastname: Williams
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry* / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D008956:Models, Chemical; D008969:Molecular Sequence Data; D020559:Monomeric GTP-Binding Proteins / Q000737:chemistry* / Q000235:genetics; D054876:Prenylation; D020539:Sequence Analysis, Protein / Q000379:methods; D012984:Software
chemical_list: D020662:Guanine Nucleotide Exchange Factors; C115955:RAP1GDS1 protein, human; D020559:Monomeric GTP-Binding Proteins
keywords: FTI; GGTI; Isoprenoid; K-Ras; Protein Complexes; Protein Farnesylation; Protein Isoprenylation; Rap1; Small GTPases; SmgGDS