Abstract
Aim:
Studies of eukaryotes have yielded 2 translation initiation mechanisms: a classical cap-dependent mechanism and a cap-independent mechanism proceeding through the internal ribosomal entry site (IRES). We hypothesized that it might be possible to identify compounds that may distinguish between cap-dependent translation and cap-independent IRES-mediated translation.
Methods:
To facilitate compound screening, we developed bicistronic reporter constructs containing a β-galactosidase gene (β-gal) and a secreted human placental alkaline phosphatase (SEAP) reporter gene. Following transcription, the β-gal gene is translated by a cap-dependent mechanism, while SEAP expression is controlled by the IRES derived from either enterovirus 71 (EV-71) or encephalomyocardi-tis virus (EMCV). This assay could potentially identify compounds that inhibit SEAP expression (cap-independent) without affecting β-gal activity (cap-dependent).
Results:
Using a bicistronic plasmid-based transient transfection assay in the COS-1 cells, we identified amantadine, a compound that inhibited the IRES of EV71- and EMCV-mediated cap-independent translation but did not interfere with cap-dependent translation when the dose of amantadine was lower than 0.25 mg/mL.
Conclusion:
These results imply that amantadine may distinguish between cap-dependent translation and cap-independent IRES-mediated translation and can be used to regulate gene expression at a translational level.
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This work was supported by Grants NSC-94-2317-B-033-001 and NSC-95-2317-B-033-001 to TYWU.
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Chen, Yj., Zeng, Sj., Hsu, J. et al. Amantadine as a regulator of internal ribosome entry site. Acta Pharmacol Sin 29, 1327–1333 (2008). https://doi.org/10.1111/j.1745-7254.2008.00876.x
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DOI: https://doi.org/10.1111/j.1745-7254.2008.00876.x
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