Transcriptional and post-transcriptional enhancement of gene expression by the 5′ UTR intron of rice rubi3 gene in transgenic rice cells

Introns play a very important role in regulating gene expression in eukaryotes. In plants, many introns enhance gene expression, and the effect of intron-mediated enhancement (IME) of gene expression is reportedly often more profound in monocots than in dicots. To further gain insight of IME in monocot plants, we quantitatively dissected the effect of the 5' UTR intron of the rice rubi3 gene at various gene expression levels in stably transformed suspension cell lines. The intron enhanced the GUS reporter gene activity in these lines by about 29-fold. Nuclear run-on experiments demonstrated a nearly twofold enhancement by the 5' UTR intron at the transcriptional level. RNA analysis by RealTime quantitative RT-PCR assays indicated the intron enhanced the steady state RNA level of the GUS reporter gene by nearly 20-fold, implying a strong role of the intron in RNA processing and/or export. The results also implicated a moderate role of the intron in enhancement at the translational level (similar to 45%). Moreover, results from a transient assay experiment using a shortened exon 1 sequence revealed an important role of exon I of rubi3 in gene expression. It may also hint a divergence in IME mechanisms between plant and animal cells. These results demonstrated transcriptional enhancement by a plant intron, but suggested that post-transcriptional event(s) be the major source of IME.