Binding of the angiogenic/senescence inducer CCN1/CYR61 to integrin alpha(6)beta(1) drives endocrine resistance in breast cancer cells

C CCN1/CYR61 promotes angiogenesis, tumor growth and chemoresistance by binding to its integrin receptor alpha(v)beta(3) in endothelial and breast cancer (BC) cells. CCN1 controls also tissue regeneration by engaging its integrin receptor alpha(6)beta(1) to induce fibroblast senescence. Here, we explored if the ability of CCN1 to drive an endocrine resistance phenotype in estrogen receptor-positive BC cells relies on interactions with either alpha(v)beta(3) or alpha(6)beta(1). First, we took advantage of site-specific mutagenesis abolishing the CCN1 receptor-binding sites to alpha(v)beta(3) and alpha(6)beta(1) to determine the integrin partner responsible for CCN1-driven endocrine resistance. Second, we explored a putative nuclear role of CCN1 in regulating ER alpha-driven transcriptional responses. Retroviral forced expression of a CCN1 derivative with a single amino acid change (D125A) that abrogates binding to alpha(v)beta(3) partially phenocopied the endocrine resistance phenotype induced upon overexpression of wild-type (WT) CCN1. Forced expression of the CCN1 mutant TM, which abrogates all the T1, H1, and H2 binding sites to alpha(6)beta(1), failed to bypass the estrogen requirement for anchorage-independent growth or to promote resistance to tamoxifen. Wild-type CCN1 promoted estradiolindependent transcriptional activity of ER alpha and enhanced ER alpha agonist response to tamoxifen. The alpha(6)beta(1)-bindingdefective TM-CCN1 mutant lost the ER alpha co-activator-like behavior of WT-CCN1. Co-immunoprecipitation assays revealed a direct interaction between endogenous CCN1 and ER alpha, and in vitro approaches confirmed the ability of recombinant CCN1 to bind ER alpha. CCN1 signaling via alpha(6)beta(1), but not via alpha(v)beta(3), drives an endocrine resistance phenotype that involves a direct binding of CCN1 to ER alpha to regulate its transcriptional activity in ER+ BC cells.