Short-interfering RNAs (siRNAs) are sequence-specific post-transcriptional regulators of gene expression resulting in degradation of homologous messenger RNA (mRNA). It is widely assumed that the events that follow mRNA cleavage involves the rapid degradation of the remaining mRNA fragments which renders the transcription-translation incompetent. However, the molecular pathways involved in post-cleavage events and mRNA decay processes are not fully understood. By using specific siRNAs and short-hairpin RNAs (shRNAs), we demonstrated that the 5’ and 3’ mRNA post-cleavage fragments of human papillomavirus type 16 (HPV-16) E7 mRNA, which plays a key role in the carcinogenesis of cervix, are unevenly degraded. Interestingly, the 5’ end of mRNA fragment was more stable and abundant than its corresponding 3’ end mRNA fragment in the RNAi-treated cells. This was confirmed by tagging E7 gene (FLAG tag at 5’ end and MYC tag at 3’ end) and treated cells were western blotted. The results showed that the C-terminally truncated proteins were actively translated and produced in treated cells.
Generally, these results provide a new understanding about the degradation of siRNA-targeted transcripts and demonstrate that RNAi can modify protein expression in cells owing to preferential stabilization and translation of the 5’ cleavage fragment. These findings question the current model of siRNA-mediated RNAi and represent a significant step forward towards understanding non-canonical pathways of siRNA targeted gene-silencing.