Abstract 156: LncRNA BC200 is induced by estradiol and regulates apoptosis in MCF-7 breast cancer cells

Abstract

Abstract LncRNA BC200 is a small functional RNA which has been implicated in the regulation of protein synthesis in neurons. It is dysregulated in invasive carcinomas of the breast. In this study, we reported that BC200 is upregulated in breast cancer, as detected by RT-PCR analysis of breast cancer tissue scan, suggesting its possible role as an oncogene. We then demonstrated that BC200 is transcriptionally induced by estradiol (E2) treatment in ER positive breast cancer cell lines through multiple functional estrogen response elements (EREs), including the full ERE sequence ∼800bp from transcriptional start site. Luciferase reporter assay combined with mutation analysis indicated that the full ERE region is critical to the estradiol-induced promoter activity. Furthermore, BC200 is crucial for cell growth and viability; its knockdown induces apoptosis in MCF-7 cells. This may explain in part why estrogen can promote cell proliferation in ER positive breast cancer cells. To better study its role in breast cancer development and progression, we generated BC200 knockout MCF-7 cell cells using CRISPR/Cas9 technique. BC200 knockout leads to the release of cytochrome c from mitochondria and promotes the cleavage of PARP-1, both of which are an indicator of cell apoptosis. This was further confirmed by TUNEL assay. It has been reported that hnRNP A2/B1 alters the bcl-X pre-mRNA splicing in favor of anti-apoptotic bcl-XL. Of great interest, BC200 knockout enhances expression of pro-apoptotic bcl-XS form at both mRNA and protein level. RNA pulldown assays using biotin-labeled BC200 probe suggested that BC200 can bind to hnRNP Q and hnRNP A2/B1. On the other hand, RNA immunoprecipitation using hnRNP A2/B1 antibody in BC200 knockout MCF-7 cells showed a significant reduction in enrichment of bcl-X mRNA as compared to control cells. We propose that the regulation of bcl-X alternative splicing depends upon endogenous expression of BC200 that would help hnRNP A2/B1 to position itself on bcl-X mRNA and modulate splicing in favor of bcl-XL. Thus, loss of BC200 leads to generation of bcl-XS protein synthesis and to apoptosis. Together, these results suggest that BC200 functions as a negative regulator of apoptosis in breast cancer and thus, it may serve as a potential target for cancer therapies. Citation Format: Ramesh Singh, Yin-yuan Mo. LncRNA BC200 is induced by estradiol and regulates apoptosis in MCF-7 breast cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 156. doi:10.1158/1538-7445.AM2015-156