THE 2 GUANINE-NUCLEOTIDE EXCHANGE FACTOR DOMAINS OF TRIO LINK THE RAC1 AND THE RHOA PATHWAYS IN-VIVO

Trio contains two functional guanine nucleotide exchange factors (GEF) domains for the Rho-like GTPases and a serine/threonine kinase domain . In vitro, GEF domain 1(GEFD1) is specifically active on Rac1, while GEF domain 2 (GEFD2) targets RhoA. To determine whether Trio could act ivate Rad and RhoA in vivo, we measured the effect of Trio on Mitogen Activated Protein Kinase (MAPK) pathways and cytoskeletal rearrangment s events mediated by the two GTPases. We show that: (i) the GEFD1 doma in of Trio triggers the MAPK pathway leading to Jun kinase (JNK) activ ation and the production of membrane ruffles; (ii) co-expression of th e TrioGEFD1 domain with a dominant-negative form of Rac blocked JNK in duction, whereas a dominant-negative form of Cdc42 did not; (iii) a de letion mutant of TrioGEFD1 lacking a region important for exchange act ivity could not stimulate JNK activity; (iv) in contrast, the TrioGEFD 2 domain does not stimulate JNK activity and induces the formation of stress fibers, as does activated RhoA; (v) furthermore, co-expression of both GEF domains induces simultaneously the formation of ruffles an d stress fibers. Trio, therefore represents a unique member of the Rho -GEFs family possessing two functional domains of distinct specificiti es, that allow it to link Rho and Rac signaling pathway in vivo.