Abstract 28: Deregulation of NAC complex inhibits muscle differentiation and blocks apoptosis in rhabdomyosarcoma cells

Abstract

Abstract Rhabdomyosarcoma (RMS) is a common malignant soft tissue sarcoma in children with limited treatment options and high failure rate during standard therapy. New therapeutic targets or strategies are therefore in high demand. The NAC complex performs many diverse biological functions, and the deregulation of its subunits has been correlated with many cancers. We sought to understand the function of NAC complex in normal muscle cells and rhabdomyosarcoma cells. With western blot, quantitative real time PCR (qRT-PCR), chromatin immunoprecipitation and immunostaining analysis, we found that the muscle specific subunit of NAC complex, skNAC, which is the alternatively spliced isoform of NACα, was downregulated in RMS cells. In normal cells, skNAC shuttled from the cytoplasm to the nucleus upon differentiation. We also showed that skNAC associated with muscle specific promoters together with BTF3 in differentiated cells, and this association is dependent on the expression of BTF3. The expression of skNAC was regulated by the splicing factor SRPK3, a serine arginine protein kinase. Overexpression of SPRK3 induced skNAC expression and muscle differentiation. We also detected the deregulation of another NAC complex subunit, NACβ (also known as BTF3). We found that BTF3 is highly overexpressed and localized in the nucleus in both RMS cell lines and human RMS tumor samples compared with normal samples. Depletion of BTF3 induced apoptosis as detected by the strong cleavage of PARP and caspase 3. BTF3 depletion also decreased the cell viability in RMS by a cologenic assay. We also confirmed that BTF3 downregulation promoted muscle gene expression and induced muscle differentiation in RMS cells. However, BTF3 played a different role in normal cells. With the deletion of BTF3 in C2C12 cells, no apoptosis signal was detected and cell differentiation was inhibited, which suggested that a precise level of BTF3 is important for normal cell function. In conclusion, we found that the NAC complex subunits, skNAC and BTF3, were deregulated in RMS cells. The restoration of skNAC by SRPK3 overexpression rescued muscle differentiation. BTF3 functioned as an anti-apoptosis factor in RMS cells and depletion of BTF3 induced apoptosis and promoted differentiation. With no apoptosis effect in normal cells, BTF3 could serve as a cancer specific therapeutic target in RMS cells. Citation Format: Meiling Zhang, Judith Davie. Deregulation of NAC complex inhibits muscle differentiation and blocks apoptosis in rhabdomyosarcoma 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 28. doi:10.1158/1538-7445.AM2015-28