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

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Abstract

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 (∼45%). Moreover, results from a transient assay experiment using a shortened exon 1 sequence revealed an important role of exon 1 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.

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Acknowledgments

The project was, in part, supported by grants from the Consortium for Plant Biotechnology Research, Inc. (St. Simons Island, GA, USA, GO12026-162 and GO12026-171 to R.Q.) and Syngenta Biotechnology Inc. (Research Triangle Park, NC, USA). The authors thank Dr. J. Nicholson for technical advice on nuclear run-on assay, Drs. L. Mathies and F. Corbin for using their facilities, and M. Massel for technical assistance. We also like to thank Drs. R. Wu and M. Ryba-White for providing rice Act1 gene clones. The authors are grateful to Plant Analysis Group at Syngenta Biotechnology Inc. for performing the RealTime PCR assays to determine transgene copy number, and to Dr. R. Shi for technical suggestions in the RealTime qRT-PCR assays. Our gratitude also extends to Dr. R. S. Boston for helpful discussion and critical reading of the manuscript.

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  1. Partha Samadder

    Present address: Department of Immunobiology, College of Medicine, University of Arizona, Tucson, AZ, 85724, USA

  2. Jianli Lu

    Present address: Department of Plant Biology, North Carolina State University, 4210 Gardner Hall, P.O. Box 7612, Raleigh, NC, 27695, USA

Authors and Affiliations

  1. Department of Crop Science, North Carolina State University, 1126A Williams Hall, P.O. Box 7620, Raleigh, NC, 27695, USA

    Partha Samadder, Elumalai Sivamani, Jianli Lu & Rongda Qu

  2. Syngenta Biotechnology, Inc., Research Triangle Park, Raleigh, NC, 27709, USA

    Xianggan Li

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  1. Partha Samadder
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Correspondence to Rongda Qu.

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Communicated by Akhilesh Tyagi.

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Samadder, P., Sivamani, E., Lu, J. et al. Transcriptional and post-transcriptional enhancement of gene expression by the 5′ UTR intron of rice rubi3 gene in transgenic rice cells. Mol Genet Genomics 279, 429–439 (2008). https://doi.org/10.1007/s00438-008-0323-8

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