Function and regulation of the Rho guanine nucleotide exchange factor Trio

Abstract

Rho GTPases oscillate between an inactive GDP-bound state and an active GTP-bound state. They are activated by Rho Guanine nucleotide Exchange Factors (GEF), which accelerate the GDP to GTP exchange. RhoGEFs fall into two different classes: the Dbl family and the DOCK family of proteins. In this review, we focus on the function and regulation of the Dbl family RhoGEF Trio. Trio and its paralog Kalirin are unique within this family in that they display two GEF domains of distinct specificity. Trio is a major regulator of neuronal development, and its function is conserved through evolution. Moreover, Trio plays an important role in cell adhesion and in signaling pathways elicited by Gαq protein-coupled receptors. Combined, these observations suggest that Trio has a major role in cellular physiology. Of note, Trio is an essential gene for mouse development, with a prominent role in the development of the nervous system. Finally, Trio expression is significantly increased in different types of tumors and it has been proposed that it could participate in oncogenesis.

Keywords: GEF; GEF inhibitors; Rho GTPases; Trio; UNC-73; actin cytoskeleton remodeling; cancer; cell adhesion; migration and invasion; neuronal differentiation.

Figure 1. Regulation of RhoGTPase activity by three classes of proteins: GEFs, GAPs, and GDIs. Rho GTPases are basic switches that oscillate between an inactive GDP-bound state and an active GTP-bound state. The family of GDIs sequesters the GTPases in the cytosol before they are targeted to the plasma membrane where they are activated by GEFs. GEFs, which receive activation by upstream signals, promote GDP/GTP exchange on the GTPases. Once bound to GTP, GTPases specifically bind to numerous effectors that trigger various cellular responses. GAPs catalyze the hydrolysis of GTP, rendering the GTPase inactive.

Figure 2. The family of Trio proteins. The name of Trio refers to the fact that Trio displays three enzymatic domains, two GEF and one serine kinase domains. In addition, Trio harbors numerous accessory domains. Listed from N-terminus to C-terminus, these include: a CRAL-Trio/Sec14 motif, several spectrin-like repeats, two SH3 motifs and an Immunoglobulin (Ig) domain. The Trio family consists of two paralogs in mammals, Trio and Kalirin. Two orthologs of Trio/Kalirin exist in invertebrates, UNC-73 in C. elegans and D-Trio in D. melanogaster. The overall organization of Trio family proteins in different organisms is remarkably conserved. The main divergence between the proteins occurs after the second GEF domain, where only vertebrate Trio and Kalirin proteins harbor a serine kinase domain. The percentage of identity at the protein level between Trio and the other members is shown.

Figure 3. Trio isoforms. In addition to its complex structure, the trio gene can encode several isoforms as a result of alternative splicing. Five isoforms (TrioA to E) that differ in their C-terminus have been described to be specific of the nervous system, with TrioC/Solo expressed only after birth in the Purkinje cells of cerebellum. In addition, a sixth, oncogenic isoform of Trio, named Tgat, has been isolated from ATL (adult-T cell leukemia) patients.

Figure 4. Trio and upstream regulators in axon outgrowth and guidance. (A) Trio mediates Rac1 activation and axon outgrowth and guidance in response to netrin-1 in mammals. Phosphorylation of Trio by the Src-family tyrosine kinase Fyn is essential for Rac1 activation by netrin-1 and for the proper targeting of DCC to the cell surface of growth cones in order to mediate netrin-1-induced cortical axon outgrowth. (B) In C. elegans, UNC-73 acts upstream of UNC-40 (ortholog of DCC) and of SAX-3 (ortholog of Robo) to positively regulate their levels at the cell membrane. UNC-73 cooperates positively with the kinesin-like protein VAB-8 and, in the SAX-3 pathway, negatively with CRML-1, the C. elegans ortholog of CARMIL (capping protein, Arp2/3, and Myosin-I linker), a protein controlling actin dynamics in lamellipodia in mammals.

Figure 5. Trio in cell adhesion. (A) Trio is activated by binding to adhesive receptors, such as cadherins and integrins of the ICAM family and thereby participates to several cell adhesion-mediated physiological processes, mainly through Rac1 activation and actin cytoskeleton remodeling. (B) Trio impinges on adhesive receptor expression at the transcriptional level through Rac1 activation. Left panel: Trio-mediated Rac1 signaling induces the phosphorylation of the transcriptional repressor Tbx3, leading to a decrease in E-cadherin expression. Tara upregulates E-cadherin transcription, by inhibiting Trio. Right panel: Trio regulates a novel pro-inflammatory pathway during transendothelial migration by controlling the expression of VCAM-1/ICAM-1 through Rac1-mediated activation of the transcription factor Ets2 in response to the cytokine TNFα.

Figure 6. Trio in physiological processes initiated by Gαq-protein coupled receptors. In vitro binding of Gαq to Trio releases the PH2-mediated inhibition of DH2, and thereby activates the GEFD2 activity toward RhoA. (A) In C. elegans, EGL-30/Gαq binding to UNC-73 isoforms activates RhoA, leading to egg laying, locomotion and growth of the animal together with the EGL-8/PLCβ pathway. UNC-73 isoforms are therefore major effectors of the Gαq pathway and play a major role in the development and the physiology of the worm. (B) In mammals, Trio mediates the Gαq-elicited mitogenic response through Rac1 and RhoA activation, by acting on p38 and Jun kinase (JNK), which control nuclear AP-1 activity.