Browsing by Author "Barbacid, Mariano"
Now showing 1 - 3 of 3
- Results Per Page
- Sort Options
Item Cbl-b, a member of the Sli-1/c-Cbl protein family, inhibits Vav-mediated c-Jun N-terminal kinase activation.(Oncogene, 1997) Bustelo, Xose R; Crespo, Piero; López Barahona, Mónica; Gutkind, J Silvio; Barbacid, MarianoWe have used the yeast two-hybrid system to identify proteins that interact with Vav, a GDP/GTP exchange factor for the Rac-1 GTPase that plays an important role in cell signaling and oncogenic transformation. This experimental approach resulted in the isolation of Cbl-b, a signal transduction molecule highly related to the mammalian c-cbl proto-oncogene product and to the C. elegans Sli-1 protein, a negative regulator of the EGF-receptor-like Let23 protein. The interaction between Vav and Cbl-b requires the entire SH3-SH2-SH3 carboxy-terminal domain of Vav and a long stretch of proline-rich sequences present in the central region of Cbl-b. Stimulation of quiescent rodent fibroblasts with either epidermal or platelet-derived growth factors induces an increased affinity of Vav for Cbl-b and results in the subsequent formation of a Vav-dependent trimeric complex with the ligand-stimulated tyrosine kinase receptors. During this process, Vav, but not Cbl-b, becomes highly phosphorylated on tyrosine residues. Overexpression of Cbl-b inhibits the signal transduction pathway of Vav that leads to the stimulation of c-Jun N-terminal kinase. By contrast, expression of truncated Cbl-b proteins and of missense mutants analogous to those found in inactive Sli-1 proteins have no detectable effect on Vav activity. These results indicate that Vav and Cbl-b act coordinately in the first steps of tyrosine protein kinase receptor-mediated signaling and suggest that members of the Sli-1/Cbl family are also negative regulators of signal transduction in mammalian cells.Item Rac-1 dependent stimulation of the JNK/SAPK signaling pathway by Vav.(Oncogene, 1996) Crespo, Piero; Bustelo, Xose R; Aaronson, David S; Coso, Ornar A; López Barahona, Mónica; Barbacid, Mariano; Gutkind, J SilvioThe protein product of the human vav oncogene, Vav exhibits a number of structural motifs suggestive of a role in signal transduction pathways, including a leucine-rich region, a plekstrin homology (PH) domain, a cysteine-rich domain, two SH3 regions, an SH2 domain, and a central Dbl homology (DH) domain. However, the transforming pathway(s) activated by Vav has not yet been elucidated. Interestingly, DH domains are frequently found in guanine nucleotide-exchange factors for small GTP-binding proteins of the Ras and Rho families, and it has been recently shown that, whereas Ras controls the activation of mitogen activated kinases (MAPKs), two members of the Rho family of small GTPases, Rac 1 and Cdc42, regulate activity of stress activated protein kinases (SAPKs), also termed c-jun N-terminal kinases (JNKs). The structural similarity between Vav and other guanine nucleotide exchange factors for small GTP-binding proteins, together with the recent identification of biochemical routes specific for members of the Ras and Rho family of GTPases, prompted us to explore whether MAPK or JNK are downstream components of the Vav signaling pathways. Using the COS-7 cell transient expression system, we have found that neither Vav nor the product of the vav proto-oncogene, proto-Vav, can enhance the enzymatic activity of a coexpressed, epitope tagged MAPK. On the other hand, we have observed that, whereas proto-Vav can slightly elevate JNK/SAPK activity, oncogenic Vav potently activates JNK/SAPK to an extent comparable to that elicited by two guanine-nucleotide exchange factors for Rho family members, Dbl and Ost. We also show that point mutations in conserved residues within the cysteine rich and DH domains of Vav both prevent its ability to activate JNK/SAPK and render Vav oncogenically inactive. In addition, we found that coexpression of the Rac-1 N17 dominant inhibitory mutant dramatically diminishes JNK/SAPK stimulation by Vav, as well as reduces the focus-forming ability of Vav in NIH3T3 murine fibroblasts. Taken together, these findings provide the first evidence that Rac-1 and JNK are integral components of the Vav signaling pathway.Item The TC21 oncoprotein interacts with the Ral guanosine nucleotide dissociation factor.(Oncogene, 1996) López Barahona, Mónica; Bustelo, Xose R; Barbacid, MarianoTC21 is a highly oncogenic member of he Ras superfamily of small GTP binding proteins. We have used the yeast two hybrid system to identify proteins that interact with an oncogenic form of the TC2I protein. cDNA clones encoding the carboxy-terminal region of the RalGDS protein were isolated from human B-cell and HeLa cDNA libraries. RalGDS is an exchange factor that stimulates GDP dissociation from Ral, another member of the Ras superfamily of proteins. The interaction between RalGDS to TC21 is direct and appears to be mediated by the effector domain of TC21 and the carboxy-terminal region of RalGDS. Moreover, RalGDS only binds to TC21 in its active, GTP-loaded configuration. These results suggest that RalGDS might be an effector molecule for TC21 and may participate in cross-talking between Ral and TC2I signalling pathways.