Poster: Phytoremediation
Abs #
985: Application of Internal Entry Sites (IRESs)-Mediated Polycistron on Co-ordinate Overexpression of Trans-genes in Plant
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Presenter: |
Li, Yujing , yujingli@arches.uga.edu |
Authors | Li, Yujing (A) Rahman, Rahman A (A) Balish, Rebacca S (A) Meagher, Richard B (A) | | Affiliations: |
(A): The University of Georgia
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Metabolic engineering requires manipulating and overexpressing numerous genes simultaneously. Polycistronic mRNAs expressing multiple proteins could be a powerful aid to this process by increasing the economy of promoter use, avoiding the negative impact of position effects and gene silencing. However, while policystronic mRNAs are common in bacteria, they are rare in eukaryotes. Internal ribosome entry sites sequences (IRESs) initiate translation without the aid of a 5'terminal cap structure. IRESs are found within the 3'UTR of many euharyotic mRNAs, including transcripts from viral and nuclear genomes. To explore if IRESs previously identified in the genomes of viruses, animals and humans can drive the translation of polycistrons in plants, dicistronic mRNAs were used to overexpress MerB and MerA proteins. MerB and MerA catalyze the reduction of organic mercury to Hg(0) and Hg(II) to Hg(0), respectively, in aerobic bacteria. The dicistrons were made with the constitutive actin2 promoter (ACT2p), merB in the first cistronic position and merA in the second cistronic position.Two IRESs tested in this study, CrTMV, an IRES from the crucifer tobacco mosaic virus genome, and eIF4G, an IRES from a human nuclear gene for a translational initiation factor, successfully drove high level expression of MerA located in the second cistronic position. The MerA protein levels from these two dicistronic constructs were 50% and 70%, respectively, of those levels obtained by direct expression of MerA alone from the ACT2p. Metal resistance assay on plants showed dramatic increases in resistance to both Hg(II) and organic mercury (PMA) as compared to WT. These data suggest there is great potential for IRES-mediated polycistronic mRNAs for the coexpression of multiple genes in plants.
