α6β1 integrin induces proteasome-mediated cleavage of erbB2 in breast cancer cells

ErbB2 and α 6 integrin have been implicated in malignancy of breast cancer cells. Here we have determined the influence of α 6 β 1 integrin on erbB2 signaling in anchorage-independent growth, using MDA-MB435 breast cancer cells. Firstly, we transfected the cells with erbB2 cDNA, and isolated cells with high or low levels of α 6 β 1 integrin by cell sorting ( α 6H-ErbB and α 6L-ErbB). We found that an erbB ligand, heregulin β 1, enhanced growth activity of α 6L-ErbB cells, but not α 6H-ErbB cells. Secondly, we established cells expressing a β 4 integrin deletion mutant ( β 4-Δcyt), which selectively inhibited α 6 β 1 integrin expression and adhesion to laminin-1. Again, heregulin β 1 enhanced the growth of erbB2 cDNA-transfected β 4-Δcyt cells, but not mock cells. Western blot analysis revealed that heregulin β 1 stimulated phosphorylation of Akt and its downstream molecules, GSK3 β and p70S6kinase, and that the extent of phosphorylation was greater in ErbB2/ β 4-Δcyt cells than ErbB2/mock cells. Furthermore, we found that the erbB2 cytoplasmic domain was truncated in ErbB2/mock cells, which was independent of ligand stimulation and adhesion, and was suppressed by proteasome inhibitors. These results suggest that α 6 β 1 integrin inhibits erbB2 signals by inducing proteasome-dependent proteolytic cleavage of the erbB2 cytoplasmic domain, and may thereby contribute to the regulation of tumor growth.