| Abstract [eng] |
miRNA, siRNA, piRNA - small non-coding RNAs (sRNAs), which regulate eukaryotic biological processes by mediating translational inhibition or/and decay or cleavage of the target RNA. In plants, animals sRNAs are protected from degradation by 2'-O-methylation at their 3'-terminal nucleotides, which is carried out by a family of HEN1 proteins. Arabidopsis thaliana HEN1 modifies miRNA/miRNA*, siRNA/siRNA* duplexes. Evaluation of HEN1-binding activity with various RNA, DNA substrates showed that enzyme specifically recognises sRNA/sRNA* duplexes at the substrate binding step. Using truncated variants of HEN1 and its mutants with amino acid substitutions it is shown that both double-stranded RNA (dsRNA)-binding domains, although unequally, together with methyltransferase domain contribute to substrate binding. Based on the obtained data, a model of HEN1 interaction with sRNA/sRNA* duplex was proposed. It was determined that HEN1 peptidyl-prolyl cis-trans isomerase-like domain of previously unknown function together with a La-motif-containing and the second dsRNA-binding domains form contacts with the dsRNA-binding protein HYL1, which together with ribonuclease DCL1, zinc finger protein SERRATE (SE) perform excision of miRNA/miRNA* duplexes from predecessors at the early stage of miRNA biogenesis. After identifiction of HEN1 interaction with HYL1, DCL1 and elucidation that HEN1 doesn't form contacts with SE, a model explaining processes of late stage of miRNA biogenesis was proposed. |
