Differential effects of polyadenylation regions on gene expression in mammalian cells

Pfarr et al. (1986). DNA DOI: 10.1089/dna.1986.5.115 Citations: 74

Parts used

bGH PolyA SignalSV40 Late PolyA Signal

Key findings

  • First systematic comparison of different polyadenylation signals (bGH, SV40 early, human collagen) showing that bGH polyA reproducibly results in 3-fold higher gene expression than SV40 early or collagen polyA signals in mammalian cells. Demonstrated that presence of polyA region is essential for efficient expression and that choice of polyA signal significantly impacts steady-state mRNA and protein levels.

Abstract

The steady-state level attained for any protein in mammalian cells is in part determined by its steady-state level of mRNA. Sequence information in and around the 3' end of an RNA which is involved in specifying and regulating polyadenylation [poly(A)] may have important consequences on mRNA levels, and ultimately on expression of the protein product. In this report we compare the effects on gene expression which result from placing several different poly(A) regions, or no poly(A) region, downstream from a marker gene (galactokinase or galK) that can be readily assayed in mammalian cells. Our results demonstrate that the presence of a poly(A) region is important for efficient gene expression and that the use of the poly(A) region of bovine growth hormone (bGH) reproducibly results in three times higher expression than that of SV40 early or human collagen poly(A) regions. We further demonstrate that changing the promoter region on these chimeric transcription units does not change the effect of the poly(A) region. Neither does changing the assay gene, since comparison of the same poly(A) regions behind another marker gene (xanthine-guanine phosphoribosyl transferase or xgprt) leads to identical differences in expression. When we examine the levels of poly(A)+ RNA that result from each transcription unit, we find that they correlate precisely with the gene expression levels. Apparently the 3' end of an RNA is a determinant of steady-state mRNA levels and, in turn, the subsequent production of the protein product.