Abstract Endoplasmic reticulum (ER)-associated degradation (ERAD) components are highly induced in various tumors and are associated with cancer cell survival and resistance to anti-cancer treatments. The ERAD pathway is crucial for the cell to adapt to ER stress by degrading misfolded, unassembled and even some normal proteins in the ER lumen. Recent work in our group has led us to identify the ER lipid raft-associated 2 (ERLIN2, also known as SPFH2, C8ORF2) gene as one of the candidate oncogenes within the 8p11-12 amplicon in breast cancer based on statistical analysis of copy number increase and over expression. ERLIN2 protein, which localized specifically to the ER, is the key component of ERAD pathway. To elucidate the oncogenic properties of the ERAD factor ERLIN2 in human breast cancer, we established the MCF10A-ERLIN2 cell line, which stably over expresses ERLIN2 in human nontransformed mammary epithelial cells (MCF10A) using the pLenti6/V5-ERLIN2 construct. ERLIN2 over expressing cells and their respective parental cell lines were assayed for alterations in growth rates, growth factor-independent proliferation and other phenotypes expressed by cancer cells. From these experiments, we found that over expression of ERLIN2 in MCF10A cells results in growth factor-independent proliferation in insulin-like growth factor deficient media and formed large, highly-proliferative, and abnormally-structured colonies in three-dimensional morphogenesis assays. However, over expression of ERLIN2 in MCF10A cells did not result in cells with anchorage independent growth potential. To further elucidate the molecular basis that ERLIN2 is linked to the ER-stress response pathways in breast cancer cells, we knocked down the ER stress sensor IRE1α activity in the breast cancer cell lines in which ERLIN2 is amplified and over expressed. The knockdown of the IRE1α RNase activity, but not the kinase activity, significantly reduced protein levels of ERLIN2 in ERLIN2 amplified breast cancer cells. In addition, forced expression of wild-type IRE1α or spliced XBP1, the IRE1α target, led to increased expression of ERLIN2 in MCF10A cells. We also found that ERLIN2 protein was induced in response to the ER stress-inducing reagents Tunicamycin, Thapsigargin or growth factor depletion. Together, our findings indicate that ERLIN2 plays an unconventional oncogenic role through modulating ER stress response and ERAD pathways in a subset of breast cancer. Targeted inhibition of ERAD factor ERLIN2 in cancer could provide potential new avenues for therapeutic development. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5022.