RESUMEN
Colorectal cancer is the second most common cancer in females and the third most common cancer diagnosed in males (Torre et al.1). In 2012, there were about 1.4 million cases and 693,900 deaths due to colorectal cancer worldwide. It is more common in developed countries, and North America, Europe, and Australia have the highest incidence rates. In the United States, adults have a 5% chance of developing colorectal cancer (Cancer of the colon and rectum-SEER stat fact sheets2). Due to the high prevalence of colorectal cancer, understanding the mechanism underlying its initiation and progression in order to find better therapeutic agents will have a high impact in the field of oncology and may improve the treatment of other cancers with shared mechanistic properties. Aberrant Wnt/ß-catenin signaling is a characteristic feature of colorectal cancer development and is the focus of this review.
Asunto(s)
Neoplasias Colorrectales/metabolismo , Vía de Señalización Wnt , beta Catenina/metabolismo , Animales , Biomarcadores de Tumor/metabolismo , Neoplasias Colorrectales/terapia , Humanos , Mutación/genética , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Vía de Señalización Wnt/genéticaRESUMEN
Mutations in the neurofibromatosis type II (NF2) tumor suppressor predispose humans and mice to tumor development. The study of Nf2+/- mice has demonstrated an additional effect of Nf2 loss on tumor metastasis. The NF2-encoded protein, merlin, belongs to the ERM (ezrin, radixin, and moesin) family of cytoskeleton:membrane linkers. However, the molecular basis for the tumor- and metastasis- suppressing activity of merlin is unknown. We have now placed merlin in a signaling pathway downstream of the small GTPase Rac. Expression of activated Rac induces phosphorylation and decreased association of merlin with the cytoskeleton. Furthermore, merlin overexpression inhibits Rac-induced signaling in a phosphorylation-dependent manner. Finally, Nf2-/- cells exhibit characteristics of cells expressing activated alleles of Rac. These studies provide insight into the normal cellular function of merlin and how Nf2 mutation contributes to tumor initiation and progression.
Asunto(s)
Neurofibromina 2/metabolismo , Transducción de Señal/fisiología , Proteínas de Unión al GTP rac/metabolismo , Células 3T3 , Secuencia de Aminoácidos , Animales , Expresión Génica/fisiología , Ratones , Datos de Secuencia Molecular , Neurofibromina 2/genética , Fosforilación , Proteína de Unión al GTP cdc42/metabolismoRESUMEN
Rho GTPases are activated by a family of guanine nucleotide exchange factors (GEFs) known as Dbl family proteins. The structural basis for how GEFs recognize and activate Rho GTPases is presently ill defined. Here, we utilized the crystal structure of the DH/PH domains of the Rac-specific GEF Tiam1 in complex with Rac1 to determine the structural elements of Rac1 that regulate the specificity of this interaction. We show that residues in the Rac1 beta2-beta3 region are critical for Tiam1 recognition. Additionally, we determined that a single Rac1-to-Cdc42 mutation (W56F) was sufficient to abolish Rac1 sensitivity to Tiam1 and allow recognition by the Cdc42-specific DH/PH domains of Intersectin while not impairing Rac1 downstream activities. Our findings identified unique GEF specificity determinants in Rac1 and provide important insights into the mechanism of DH/PH selection of GTPase targets.
Asunto(s)
Proteínas Adaptadoras del Transporte Vesicular , Factores de Intercambio de Guanina Nucleótido/química , Factores de Intercambio de Guanina Nucleótido/metabolismo , Mapeo de Interacción de Proteínas , Proteína de Unión al GTP rac1/química , Proteína de Unión al GTP rac1/metabolismo , Células 3T3 , Sustitución de Aminoácidos/genética , Animales , Sitios de Unión , Proteínas Portadoras/química , Proteínas Portadoras/metabolismo , Transformación Celular Neoplásica , Activación Enzimática , Humanos , Ligandos , Ratones , Modelos Moleculares , Mutagénesis Sitio-Dirigida/genética , Mutación/genética , Unión Proteica , Estructura Terciaria de Proteína , Proteínas/química , Proteínas/metabolismo , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Transducción de Señal , Especificidad por Sustrato , Proteína 1 de Invasión e Inducción de Metástasis del Linfoma-T , Proteína de Unión al GTP cdc42/química , Proteína de Unión al GTP cdc42/genética , Proteína de Unión al GTP cdc42/metabolismo , Proteína de Unión al GTP rac1/genéticaRESUMEN
Normally, Rho GTPases are activated by the removal of bound GDP and the concomitant loading of GTP catalyzed by members of the Dbl family of guanine nucleotide exchange factors (GEFs). This family of GEFs invariantly contain a Dbl homology (DH) domain adjacent to a pleckstrin homology (PH) domain, and while the DH domain usually is sufficient to catalyze nucleotide exchange, possible roles for the conserved PH domain remain ambiguous. Here we demonstrate that the conserved PH domains of three distinct Dbl family proteins, intersectin, Dbs, and Tiam1, selectively bind lipid vesicles only when phosphoinositides are present. While the PH domains of intersectin and Dbs promiscuously bind several multiphosphorylated phosphoinositides, Tiam1 selectively interacts with phosphatidylinositol 3-phosphate (K(D) approximately 5-10 microm). In addition, and in contrast to recent reports, catalysis of nucleotide exchange on nonprenylated Rac1 provided by various extended portions of Tiam1 is not influenced by (a) soluble phosphoinositide head groups, (b) dibutyl versions of phosphoinositides, or (c) lipid vesicles containing phosphoinositides. Likewise, GEF activity afforded by DH/PH fragments of intersectin and Dbs are also not altered by phosphoinositide interactions. These results strongly suggest that unless all relevant components are localized to a lipid membrane surface, Dbl family GEFs generally are not intrinsically modulated by binding phosphoinositides.
Asunto(s)
Proteínas Adaptadoras del Transporte Vesicular , Proteínas Portadoras/metabolismo , Factores de Intercambio de Guanina Nucleótido/metabolismo , Fosfatidilinositoles/metabolismo , Proteínas/metabolismo , Immunoblotting , Unión Proteica , Factores de Intercambio de Guanina Nucleótido Rho , Resonancia por Plasmón de Superficie , Proteína 1 de Invasión e Inducción de Metástasis del Linfoma-TRESUMEN
The neural cell adhesion molecule L1 mediates the axon outgrowth, adhesion, and fasciculation necessary for proper development of synaptic connections. Mutations of human L1 cause an X-linked mental retardation syndrome termed CRASH (corpus callosum hypoplasia, retardation, aphasia, spastic paraplegia, and hydrocephalus), and L1 knock-out mice display defects in neuronal process extension resembling the CRASH phenotype. Little is known about the biochemical or cellular mechanism by which L1 performs neuronal functions. Here it is demonstrated that clustering of L1 with antibodies or L1 protein in rodent B35 neuroblastoma and cerebellar neuron cultures induced the phosphorylation/activation of the mitogen-activated protein kinases (MAPKs) and extracellular signal-regulated kinases 1 and 2. MAPK activation was essential for L1-dependent neurite outgrowth, because chemical inhibitors [2-(2'-amino-3'-methoxyphenyl)-oxanaphthalen-4-one and 1,4-diamino-2, 3-dicyano-1,4-bis(2-aminophenylthio)butadiene] of the MAPK kinase MEK strongly suppressed neurite outgrowth by cerebellar neurons on L1. The nonreceptor tyrosine kinase pp60(c-src) was required for L1-triggered MAPK phosphorylation, as shown in src-minus cerebellar neurons and by expression of the kinase-inactive mutant Src(K295M) in B35 neuroblastoma cells. Phosphatidylinositol 3-kinase (PI3-kinase) and the small GTPase p21(rac) were identified as signaling intermediates to MAPK by phosphoinositide and Rac-GTP assays and expression of inhibitory mutants. Antibody-induced endocytosis of L1, visualized by immunofluorescence staining and confocal microscopy of B35 cells, was blocked by expression of kinase-inactive Src(K295M) and dominant-negative dynamin(K44A) but not by inhibitors of MEK or PI3-kinase. Dynamin(K44A) also inhibited L1 antibody-triggered MAPK phosphorylation. This study supports a model in which pp60(c-src) regulates dynamin-mediated endocytosis of L1 as an essential step in MAPK-dependent neurite outgrowth on an L1 substrate.