The effect of intron location on intron-mediated enhancement of gene expression in Arabidopsis

Introns are often required for full expression of genes in organisms as diverse as plants, insects, nematodes, yeast, and mammals. To explore the potential mechanisms of intron-mediated enhancement in Arabidopsis thaliana, the effect of varying the position of an intron was determined using a series of reporter gene fusions between TRYPTOPHAN BIOSYNTHESIS1 (TRP1) and GUS. Two introns that differ in the degree to which they stimulate expression were individually tested at six locations within coding sequences and two positions in the 3'-UTR. The ability of the first introns from both the TRP1 and POLYUBIQUITIN10 (UBQ10) genes to elevate mRNA accumulation in transgenic plants was found to decline with distance from the promoter, despite their being efficiently spliced from all coding sequence locations. Neither intron significantly enhanced mRNA accumulation when positioned 1.1 kb or more from the start of transcription. In addition, measurements of GUS enzyme activity revealed that both introns at all locations elevated GUS activity more than they enhanced mRNA accumulation. The stimulation mediated by two of four other introns tested at the position nearest the promoter was also greater at the level of GUS activity than mRNA accumulation. These findings support a model in which introns increase transcription and promote translation by two distinct mechanisms.