Homo- and hetero-oligomerization of PDZ-RhoGEF, LARG and p115RhoGEF by their C-terminal region regulates their in vivo Rho GEF activity and transforming potential

Abstract

PDZ-RhoGEF, LARG, and p115RhoGEF are members of a newly identified family of Rho-guanine nucleotide exchange factors (GEFs) exhibiting a unique structural feature consisting of the presence of an area of similarity to regulators of G protein signaling (RGS). This RGS-like (RGL) domain provides a functional motif by which Ga 12 and Ga 13 can bind and regulate the activity of these RhoGEFs, thus providing a direct link from these heterotrimeric G proteins to Rho. PDZ-RhoGEF and LARG can also be phosphorylated by tyrosine kinases, including FAK, and associate with Plexin B, a semaphorin receptor, which controls axon guidance during development, through their PDZ domain, thereby stimulating Rho. Interestingly, while characterizing a PDZ-RhoGEF antiserum, we found that a transfected PDZ-RhoGEF construct associated with the endogenous PDZ-RhoGEF. Indeed, we observed that PDZ-RhoGEF and LARG can form homo-and hetero-oligomers, whereas p115RhoGEF can only homo-oligomerize, and that this intermolecular interaction was mediated by their unique C-terminal regions. Deletion of the C-terminal tail of PDZ-RhoGEF had no significant effect on the GEF catalytic activity towards Rho in vitro, but resulted in a drastic increase in the ability to stimulate a serum response element reporter and the accumulation of the GTP-bound Rho in vivo. Furthermore, removal of the C-termini of each of the three RGL-containing GEFs unleashed their full transforming potential. Together, these findings suggest the existence of a novel mechanism controlling the activity of PDZRhoGEF, LARG, and p115RhoGEF, which involves homo-and hetero-oligomerization through their inhibitory C-terminal region.

CD98hc (SLC3A2) regulation of skin homeostasis wanes with age

“…It was recently shown that Fyn may stimulate LARG activity downstream of integrins (Guilluy et al, 2011), but we observe, rather, an inhibition of Src family kinases upon loss of CD98hc in our experimental model, which suggests that LARG has to be activated by other means. These may possibly include phosphorylation by kinases such as PKC or FAK, for instance (Chikumi et al, 2002;Disatnik et al, 2002;Suzuki et al, 2003;Dovas et al, 2006), as well as regulation of LARG homodimerization by CD98hc, which has long been known to regulate its activity (Chikumi et al, 2004). Further molecular investigations, in vitro, in cell culture should be performed to dissect this regulation.…”

Homo-oligomerization of ALS2 through Its Unique Carboxyl-terminal Regions Is Essential for the ALS2-associated Rab5 Guanine Nucleotide Exchange Activity and Its Regulatory Function on Endosome Trafficking

“…Inhibition of oligomerization diminishes the in vivo GEF activity of Dbl (38) and abolishes ␤1PIX function in vivo (37). In the cases of p115RhoGEF, LARG, and PDZ-RhoGEF, deletion of the C-terminal parts that were required for oligomerization had no significant effect on the in vitro GEF activities but resulted in the drastic stimulation of in vivo functions (40,41). Taken together, these findings suggest that oligomerization may be an important common molecular feature in the GDP/ GTP exchanging reactions on the small GTPase, which are mediated by either some members of Rab5GEFs or RhoGEFs, albeit with fundamental differences in their physiological roles.…”

“…These findings fuel speculation that the Rab5GEFs might function as a multiple protein complex in vivo, regardless of whether homo-or heterophilic complexes are formed. By contrast, several RhoGEFs, including ␤1PIX (36, 37), Dbl (38), RasGRF1 (39), Ras-GRF2 (39), p115RhoGEF (40,41), LARG (41), and PDZ-Rho-GEF (41), have already been shown to dimerize or oligomerize. Inhibition of oligomerization diminishes the in vivo GEF activity of Dbl (38) and abolishes ␤1PIX function in vivo (37).…”

Hyaluronan-mediated CD44 activation of RhoGTPase signaling and cytoskeleton function promotes tumor progression

“…Furthermore, overexpression of LARG in acute myeloid leukemia (AML) has been shown to exhibit growth-promoting activity [78]. A number of studies show that LARG and its transforming potential can be activated by focal adhesion kinase (FAK) phosphorylation [80] and/or hetero-oligomerization through LARG's inhibitory C-terminal region [81]. Apparently, LARG-RhoGEF activity can be regulated by many different mechanisms in addition to its previously described role in Gα subunit signaling by GPCRs.…”