RESUMEN
Depending on its cellular localization, p120 catenin (p120ctn) can participate in various processes, such as cadherin-dependent cell-cell adhesion, actin cytoskeleton remodeling, and intracellular trafficking. Recent studies also indicate that p120ctn could regulate cell proliferation and contact inhibition. This report describes a new function of p120ctn in the regulation of cell cycle progression. Overexpression of the p120ctn isoform 3A in human colon adenocarcinoma cells (HT-29) results in cytoplasmic accumulation of the protein, as observed in many tumors. This cytoplasmic increase is correlated with a reduction in proliferation and inhibition of DNA synthesis. Under these conditions, experiments on synchronized cells revealed a prolonged S phase associated with cyclin E stabilization. Both confocal microscopy and biochemical analysis showed that cyclin E and cyclin-dependent kinase 2 colocalized with p120ctn in centrosomes during mitosis. These proteins are associated in a functional complex evidenced by coimmunoprecipitation experiments and the emergence of Thr199-phosphorylated nucleophosmin/B23. Such post-translational modification of this centrosomal target has been shown to trigger the initiation of centrosome duplication. Therefore, p120ctn-mediated accumulation of cyclin E in centrosomes may participate in abnormal amplification of centrosomes and the inhibition of DNA replication, thus leading to aberrant mitosis and polyploidy. Because these modifications are often observed in cancer, p120ctn may represent a new therapeutic target for future therapy.
Asunto(s)
Moléculas de Adhesión Celular/metabolismo , Neoplasias del Colon/metabolismo , Neoplasias del Colon/patología , Ciclina E/metabolismo , Quinasa 2 Dependiente de la Ciclina/metabolismo , Fosfoproteínas/metabolismo , Cateninas , Moléculas de Adhesión Celular/biosíntesis , Moléculas de Adhesión Celular/genética , Ciclo Celular/fisiología , Procesos de Crecimiento Celular/fisiología , Centrosoma/metabolismo , Neoplasias del Colon/genética , Citoplasma/metabolismo , Progresión de la Enfermedad , Amplificación de Genes , Inestabilidad Genómica , Proteínas Fluorescentes Verdes/biosíntesis , Proteínas Fluorescentes Verdes/genética , Células HT29 , Humanos , Fosfoproteínas/biosíntesis , Fosfoproteínas/genética , Fosforilación , Proteínas Recombinantes de Fusión/biosíntesis , Proteínas Recombinantes de Fusión/genética , Regulación hacia Arriba , Catenina deltaRESUMEN
Human intestinal cell differentiation is mediated by signaling pathways that remain largely undefined. We and others have shown that cell migration and differentiation along the crypt-villus axis is associated with temporal and spatial modulations of the repertoire, as well as with the function of integrins and E-cadherins and their substrates. Cross-talk between integrin and cadherin signaling was previously described and seems to coordinate this differentiation process. Here, we report that engagement of alpha6 and, to a lesser extent, alpha3 integrin subunits after HT-29 cell adhesion on laminin 5 increases the expression of E-cadherin, which then organizes into nascent adherens junctions. We further identify that phosphoinositide 3-kinase (PI 3-kinase) activation plays a key role in this cross-talk. Indeed, integrin-dependent adhesion on laminin 5 stimulates PI 3-kinase activity. Immunofluorescence and immunoprecipitation experiments revealed that activated PI 3-kinase is recruited at cell-cell contacts. Using LY294002, an inhibitor of PI 3-kinase activity, we found that this activation is essential for E-cadherin connection with the cytoskeleton and for biogenesis of adherens junctions. Finally, we demonstrated that PI 3-kinase could signal through Rac1b activation to control adherens junction assembly. Our results provide a mechanistic insight into integrin-cadherin cross-talk and identify a novel role for PI 3-kinase in the establishment of adherens junctions.