The Studies On The Function And Mechanism Of The C-Abl Kinase In Integrin α_v_β 3 Mediated Melanoma Cell Adhesion And Migration

Tumor cell invasion and metastasis is the main reason for the high cancer mortality rate, and cell invasion and metastasis is based on cell migration. Integrin as one of adhesion molecules on the cell membrane, one hand, it can through its intracellular domain in combination with the intracellular cytoskeleton; on the other hand through the extracellular globular domain it can link with the extracellular matrix, so that cells anchored in the extracellular matrix. In addition, integrin is critical for cell migration because integrin can integrate intracellular and extracellular signals to regulate cell migration.αvβ3 integrin in many tumor cells have high expression, including breast cancer, prostate cancer cells, melanoma cells and glioma cells.αvβ3 integrin express in tumor cells and it closely relate to tumor occurrence, especially in tumor cell invasion and metastasis.Integrin mediates adhesion and migration of tumor cells mainly through contacting with the cytoskeleton to form focal adhesion, and through regulating the dynamic temporal and spatial variation of focal adhesion assembly and disassembly. Integrin itself has no activity, its function generally regulated by a number of related kinases. c-Abl kinase as an intracellular non-receptor tyrosine kinases can phosphorylate many proteins, activate the corresponding target molecules, regulate cytoskeletal reorganization or gene expression, consequently, participate in cell apoptosis, differentiation, aging, etc. variety of physiological processes, there were studies also reported that it participates in cell migration.In the present study, we adopted transwell, monolayer-cell scratch and other experimental models of migration found that c-Abl kinase involve in the PDGF-inducedαvβ3 integrin-mediated migration of melanoma A375 cells. And by immunofluorescence double-labeling experiments confirmed the existence of two co-localized phenomenon. Immunoprecipitation experiments showed that the c-Abl kinase andαvβ3 integrin present in one protein complex. By using in vitro kinase reaction experiments, we found that c-Abl kinase interactions withαvβ3 integrin dependent on c-Abl kinase activity changes. while c-Abl kinase in the regulation ofαvβ3 integrin is independent of F-actin effect and c-Abl kinase is not involved inαvβ3 integrin and extracellular ligand-induced integrin outside-in signaling pathways.We constructed several different interactions with other proteins c-Abl kinase critical domain GST fusion protein, by using in vitro expression system GST-pull down experiments we found that c-Abl kinase through its SH3 domain interact withαvβ3 integrin to form complex. We further confirm that c-Abl protein kinase is a direct connection with the talin to regulateαvβ3 integrin functions in cell migration. By point mutation experiments, we found that the 89 and 134 tyrosine of c-Abl kinase SH3 domains for the combination of c-Abl and talin are not the only key point of its role. These studies reveal the mechanism of melanoma A375 cell migration and also provides a reliable data for future research.