The internal ribosome entry site (IRES) enables polycistronic expression of multiple genes from a single mRNA controlled by one promoter. In general, only the most 5′ coding sequence is abundantly translated from polycistronic mRNAs in eukaryotic cells; IRES-mediated translation allows additional coding sequences at other positions to be translated at detectable levels. However, IRES-mediated translation often results in much lower protein expression than translation of single-gene mRNAs. We first aimed to improve IRES-mediated gene expression with a transient expression system based on
Nicotiana benthamiana. We demonstrated that the presence of two cassettes comprised of an IRES-mediated Venus coding sequence results in higher Venus expression than one cassette. The double IRES cassette system is expected to be useful for expressing a gene of interest polycistronically in plants. Using the double IRES cassette system, we found that polycistronic expression of a reporter gene and two copies of an IRES-mediated RNA silencing suppressor gene protected the transcribed mRNA from RNA silencing-mediated degradation. This is the first report of a mRNA self-protection system from RNA
silencing by IRES-mediated expression of a viral suppressor in plants.