The non-coding RNA BC200 associates with polysomes to positively regulate mRNA translation in tumour cells.

BC200 is a non-coding RNA elevated in a broad spectrum of tumour cells that is critical for cell viability, invasion, and migration. Over-expression studies have implicated BC200 and the rodent analog BC1 as negative regulators of translation in both cell-based and in vitro translation assays. While consistent, these studies have not been confirmed in knock-down studies and direct evidence for this function is lacking. Herein, we have demonstrated that BC200 knock-down is correlated with a decrease in global translation rates. As this conflicts with the hypothesis that BC200 is a translational suppressor, we overexpressed BC200 by transfection of in vitro transcribed RNA and transient expression from transfected plasmids. In this context BC200 suppressed translation; however, an innate immune response confounded the data. To overcome this, breast cancer cells stably overexpressing BC200 and various control RNAs were developed by selection for genomic incorporation of a plasmid co-expressing BC200 and the neomycin resistance gene. Stable overexpression of BC200 was associated with elevated translation levels in pooled stable cell lines and isolated single-cell clones. Crosslinking sucrose density gradient centrifugation demonstrated an association of BC200 and its reported binding partners SRP9/14, CSDE1, DHX36 and PABPC1 with both ribosomal subunits and polysomal RNA, an association not previously observed due to the labile nature of the interactions. In summary, these data present a novel understanding of BC200 function as well as optimized methodology that has far reaching implications in the study of non-coding RNAs, particularly within the context of translational regulatory mechanisms.