Effects of 3′-untranslated region modifications on transgene expression in the human embryonic kidney 293 cell line

Abstract

Improving the productivity of recombinant cell lines is a key goal of mammalian cell line development (CLD), which can be achieved via different strategies, such as the control of mRNA degradation. However, such strategies have received little attention. Moreover, only a few studies have attempted to use the 3′-untranslated region (UTR) to alter the mRNA stability and protein expression levels in CLD. Whether this strategy can help in controlling the transgene expression remains unclear. Therefore, to determine the utility of 3′-UTR engineering for CLD, we aimed to assess whether mRNA stability and transgene expression can be altered by inserting additional 3′- UTR sequences into the commonly used expression cassette in the representative human cell line, human embryonic kidney (HEK) 293. We designed synthetic 3′- UTRs composed of various motifs derived from endogenous genes based on global mRNA stability and transcriptome data of HEK293 cells. Although changes in the mRNA half-life did not have any direct effects, the inserted 3′-UTR sequences exhibited potential to modulate the mRNA half-life and protein expression levels. These results demonstrate the potential use of 3′-UTR engineering for the fine-tuning of transgene expression and tight regulation of gene of interest in mammalian cell line engineering.