Casitas B-cell lymphoma (Cbl) proteins protect mammary epithelial cells from proteotoxicity of active c-Src accumulation

Casitas B-cell lymphoma (Cbl) family ubiquitin ligases negatively regulate tyrosine kinase-dependent signal transduction by promoting degradation of active kinases. We and others previously reported that loss of Cbl functions caused hyperproliferation in lymphoid and hematopoietic systems. Unexpectedly, Cbl deletion in Cbl-b-null, Cbl-c-null primary mouse mammary epithelial cells (MECs) (Cbl triple-deficiency) induced rapid cell death despite enhanced MAP kinase and AKT activation. Acute Cbl triple-deficiency elicited distinct transcriptional and biochemical responses with partial overlap with previously described cellular reactions to unfolded proteins and oxidative stress. Although the levels of reactive oxygen species were comparable, detergent-insoluble protein aggregates containing phosphorylated c-Src accumulated in Cbl triple-deficient MECs. Treatment with a broad-spectrum kinase inhibitor dasatinib blocked protein aggregate accumulation and restored in vitro organoid formation. This effect is most likely mediated through c-Src because Cbl triple-deficient MECs were able to form organoids upon shRNA-mediated c-Src knockdown. Taking these data together, the present study demonstrates that Cbl family proteins are required to protect MECs from proteotoxic stress-induced cell death by promoting turnover of active c-Src.