The invasion of glioblastoma stem cells is reduced by the sequence of the connexin43 that interacts with c-Src

Abstract

Glioblastoma stem cells (GSC) constitute a niche of cells with self-renewal ability and resistance to conventional therapies. Thus, they are responsible for relapse in these aggressive brain tumors when their resection is not complete. The expression of connexin43 (Cx43), the main gap junction channel-forming protein, and c-Src, a non receptor tyrosine kinase, is inversely related in glioblastoma cells. Interestingly, restoring Cx43 in GSC reverses GSC phenotype. The sequence responsible for this effect is the region that interacts with c-Src (amino acids 266-283). The Focal Adhesion Kinase (FAK) requires the c-Src-mediated phosphorylation in tyrosines 576 and 577 in order to be fully active and promote cell migration. In this study, fresh human tumour biopsies were disaggregated to obtain human primary GSCs. These cells were treated with a cell-penetrating peptide containing the Cx43-Src interacting sequence (Tat-Cx43266-283). The migration was analyzed by tracking individual cell trajectories in cultures recorded by Time-Lapse Live-cell Imaging. Matrigel-treated transwell inserts were used to analyze the invasion and the levels of phosphorylation of FAK tyrosines 576 and 577 were evaluated by western Blot. Our results showed that Tat-Cx43266-283 reduced the migration and invasion of human primary GSC. The analysis of c-Src and FAK proteins suggests that the mechanism by which this reduction takes place includes the inhibition of c-Src/FAK axis. In conclusion, Tat-Cx43266-283 inhibits c-Src with the subsequent reduction in FAK phosphorylation required to establish appropriate focal adhesions for migration. All together, these results indicate an important antivasive role of the sequence of Cx43 that interacts with c-Src in glioma stem cells.