BAS/BSCR20 The role of a Gab1-Tribbles 2 interaction in phosphoinositide-3-kinase, AKT/PKB cascade regulation and cellular morphology

Abstract

Apoptosis is a key event in atherosclerotic plaque formation. The phosphoinositide-3-kinase (PI3K) cascade is involved in many cellular activities in plaques, such as migration and cell survival. Our previous data have identified an interaction between Tribbles 2 (Trb2) and signalling adaptor molecule Grb2 associated binder protein (Gab1). The functional consequences of this interaction are unknown. Gab1 interacts with the p85 domain of PI3K to mediate downstream activation of the Akt/PKB anti-apoptotic signalling pathway. We examined whether tribbles, a new family of signalling regulators, link with PI3K to control activation of Akt and potentially inhibit apoptosis.HEK293 cells were transfected with Trb2 and mutant or wt Gab1 and formation of Trb2/Gab1 complexes was quantified using a yellow fluorescent protein-based protein fragment complementation assay. We show that overexpression of the PI3Kδ and β (catalytic) and the PI3Kα (regulatory) subunits leads to an increase in Trb2–Gab1 interaction (relative binding intensity 2.48%± SEM vs 3.9%± SEM, p<0.05, and 2.52%± SEM vs 5.3%± SEM, respectively). These data suggest that Trb2/Gab1 binding is modulated via a PI3K dependent feedback loop and raise the possibility that that Trb2 may act as a co-regulator of Gab1. In addition, we show that Gab1 has an effect on cell morphology upon PI3K cascade activation, and this morphological consequence is potentiated further in the presence of Trb2. These findings in turn suggest that the Gab1–Trb2 interaction may participate in controlling cell survival and morphology and potentially, atherosclerotic plaque development or rupture.