RESUMO
The Pix/Cool proteins are involved in the regulation of cell morphology by binding to small Rho GTPases and kinases of the Pak family. Recently, it has been shown that betaPix/Cool-1 associates with the ubiquitin ligase Cbl, which appears to be a critical step in Cdc42-mediated inhibition of epidermal-growth-factor-receptor (EGFR) ubiquitylation and downregulation. Here we show that the SH3 domain of betaPix specifically interacts with a proline-arginine motif (PxxxPR) present within the ubiquitin ligase Cbl and Pak1 kinase. Owing to targeting of the same sequence, Cbl and Pak1 compete for binding to betaPix. In this complex, Cbl mediates ubiquitylation and subsequent degradation of betaPix. Our findings reveal a double feedback loop in which the Cdc42/betaPix complex blocks Cbl's ability to downregulate EGFR, while Cbl in turn promotes degradation of betaPix in order to escape this inhibition. Such a relationship provides a mechanism to fine-tune the kinetics of RTK endocytosis and degradation depending on the pool of active Cdc42 and the duration of EGFR signaling.
Assuntos
Proteínas de Ciclo Celular/fisiologia , Regulação da Expressão Gênica , Fatores de Troca do Nucleotídeo Guanina/fisiologia , Processamento de Proteína Pós-Traducional , Proteínas Serina-Treonina Quinases/fisiologia , Proteínas Proto-Oncogênicas c-cbl/fisiologia , Proteína cdc42 de Ligação ao GTP/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/química , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Motivos de Aminoácidos , Ligação Competitiva , Neoplasias da Mama/patologia , Proteínas de Ciclo Celular/biossíntese , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/genética , Linhagem Celular/metabolismo , Linhagem Celular Tumoral/metabolismo , Endocitose , Células Epiteliais/metabolismo , Receptores ErbB/biossíntese , Receptores ErbB/fisiologia , Retroalimentação Fisiológica , Regulação Neoplásica da Expressão Gênica , Genes erbB-1 , Fatores de Troca do Nucleotídeo Guanina/biossíntese , Fatores de Troca do Nucleotídeo Guanina/química , Fatores de Troca do Nucleotídeo Guanina/genética , Humanos , Rim , Proteínas de Neoplasias/fisiologia , Ligação Proteica , Mapeamento de Interação de Proteínas , Proteínas Serina-Treonina Quinases/química , Proteínas Proto-Oncogênicas c-cbl/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-cbl/química , Proteínas Proto-Oncogênicas c-cbl/genética , Proteínas Recombinantes de Fusão/fisiologia , Fatores de Troca de Nucleotídeo Guanina Rho , Transfecção , Ubiquitina/metabolismo , Quinases Ativadas por p21 , Proteínas rac1 de Ligação ao GTP/metabolismo , Domínios de Homologia de srcRESUMO
Cbl proteins control multiple cellular processes by acting as ubiquitin ligases and multifunctional adaptor molecules. They are involved in the control of cell proliferation, differentiation and cell morphology, as well as in pathologies such as autoimmune diseases, inflammation and cancer. Here we review recent advances in understanding the role of Cbl and the importance of a growing repertoire of Cbl-interacting proteins in the regulation of signaling pathways triggered by growth factors, antigens, cell adhesion, cytokines and hormones. We also address key issues of the nature of proteins that bind Cbl in particular cells, where they are located, and how they are altered or traffic within cells upon stimulation. It is becoming obvious that temporal and spatial changes in Cbl signaling networks are essential for the control of physiological processes in a variety of cells and organs and that their deregulation can result in the development of human diseases.
Assuntos
Proteínas Oncogênicas de Retroviridae/metabolismo , Transdução de Sinais/fisiologia , Animais , Endocitose/fisiologia , Humanos , Ligases/metabolismo , Proteína Oncogênica v-cbl , Ligação Proteica , Estrutura Terciária de Proteína , Receptores Proteína Tirosina Quinases/genética , Receptores Proteína Tirosina Quinases/metabolismo , Proteínas Oncogênicas de Retroviridae/genética , Ubiquitina/metabolismoRESUMO
During migration, tumour cells interact with neighbouring neoplastic and normal host cells, and such interaction may influence their motile activity. We investigated the effect of homotypic collisions on the motile activity of two tumour cell lines, mouse melanoma B16 and rat sarcoma XC, and nontransformed human skin fibroblasts. It was found that the tumour cells show only limited motile activity when moving as single cells without contact with neighbours. At a higher density of the culture (and also at a greater number of cell to cell contacts) the activation of motility of investigated tumour cells was observed. On the other hand, the normal human skin fibroblasts showed a typical reaction of density-dependent inhibition of motility. The motile activity of tumour cells was not affected by conditioned media and was visibly dependent on a direct physical contact among colliding cells. The activation of cell movement was observed about 40-50 min after the initial contact between tumour cells. Contact-activated migration of neoplastic cells was inhibited by 50 microM verapamil (a selective voltage-gated calcium channel inhibitor) and 10 microM gadolinium chloride (a nonspecific blocker of mechanosensitive ion channels) but not by pertussis toxin. The observation that homotypic collisions among tumour cells strongly increase their motile activity suggests that contact-activated migration may play a significant role in tumour invasion and metastasis.