Splice Variants of SmgGDS Control Small GTPase Prenylation and Membrane Localization

Abstract

Ras and Rho small GTPases possessing a C-terminal polybasic region (PBR) are vital signaling proteins whose misregulation can lead to cancer. Signaling by these proteins depends on their ability to bind guanine nucleotides and their prenylation with a geranylgeranyl or farnesyl isoprenoid moiety and subsequent trafficking to cellular membranes. There is little previous evidence that cellular signals can restrain nonprenylated GTPases from entering the prenylation pathway, leading to the general belief that PBR-possessing GTPases are prenylated as soon as they are synthesized. Here, we present evidence that challenges this belief. We demonstrate that insertion of the dominant negative mutation to inhibit GDP/GTP exchange diminishes prenylation of Rap1A and RhoA, enhances prenylation of Rac1, and does not detectably alter prenylation of K-Ras. Our results indicate that the entrance and passage of these small GTPases through the prenylation pathway is regulated by two splice variants of SmgGDS, a protein that has been reported to promote GDP/GTP exchange by PBR-possessing GTPases and to be up-regulated in several forms of cancer. We show that the previously characterized 558-residue SmgGDS splice variant (SmgGDS-558) selectively associates with prenylated small GTPases and facilitates trafficking of Rap1A to the plasma membrane, whereas the less well characterized 607-residue SmgGDS splice variant (SmgGDS-607) associates with nonprenylated GTPases and regulates the entry of Rap1A, RhoA, and Rac1 into the prenylation pathway. These results indicate that guanine nucleotide exchange and interactions with SmgGDS splice variants can regulate the entrance and passage of PBR-possessing small GTPases through the prenylation pathway.

SmgGDS Is a Guanine Nucleotide Exchange Factor That Specifically Activates RhoA and RhoC

“…Interestingly, recent results from the Williams laboratory suggest that SmgGDS-558 specifically binds prenylated GTPases in cells, whereas the larger SmgGDS isoform specifically interacts with nonprenylated GTPases (39). As the majority of Rho in cells is expected to be prenylated, this may explain our observation of preferential activation of Rho in cells by SmgGDS-558.…”

The Tumor-suppressive Small GTPase DiRas1 Binds the Noncanonical Guanine Nucleotide Exchange Factor SmgGDS and Antagonizes SmgGDS Interactions with Oncogenic Small GTPases

“…There are two predominant isoforms of SmgGDS, SmgGDS-607 and SmgGDS-558, which differ by one armadillo domain (5). Recent studies have demonstrated that SmgGDS-558 plays a more significant role than SmgGDS-607 in activating RhoA in breast cancer cells, despite lower levels of endogenous SmgGDS-558 protein (6,7).…”

“…SmgGDS (Rap1GDS1) is a noncanonical GEF for RhoA and RhoC (4) and also promotes the pro-oncogenic functions of several other small GTPases with C-terminal polybasic regions (PBRs) (5). There are two predominant isoforms of SmgGDS, SmgGDS-607 and SmgGDS-558, which differ by one armadillo domain (5).…”

Control of RhoA Methylation by Carboxylesterase I

“…Regulation of the prenylation pathway has been largely unexamined, although the recent finding that entry into, and passage through, the prenylation pathway of some small GTPases can be controlled by specific SmgGDS splice variants has highlighted the potential for control of CaaX protein function via this process (28). These recent developments in the prenylation field, coupled with the genetic evidence in plants for demethylation of prenyl proteins (18,19) and our finding of dynamic control of RhoA methylation by CES1, opens new avenues of investigation into the dynamics of prenyl protein processing.…”