Extensive 3' modification of plant small RNAs is modulated by helper component-proteinase expression.
RNA silencing is an evolutionarily conserved process in eukaryotes that represses gene expression by using 21- to 24-nt guide RNAs to mediate mRNA cleavage or translational inhibition. Plants have two distinct groups of silencing-associated small RNAs (smRNAs): the micro RNAs (miRNAs) and the small interfering RNAs (siRNAs). A recent report by Yu et al. [Yu, B., Yang, Z., Li, J., Minakhina, S., Yang, M., Padgett, R. W., Steward, R. & Chen, X. (2005) Science 307, 932-935] has shown that plant miRNAs are modified at their 3' termini with a methyl group. Here, we show that a large fraction of all silencing-associated smRNAs in tobacco are modified; this modification occurs on the 2' hydroxyl of the terminal ribose and significantly reduces the cloning efficiency of these modified smRNAs. Expression of the strong silencing suppressor P1/helper-component proteinase results in a marked decrease in the 3'-terminal modification of viral siRNAs but does not significantly affect the modification of endogenous miRNAs and 24-nt siRNAs. The differential modification mediated by helper-component proteinase expression implies that exogenous and endogenous smRNAs are processed through independent pathways that are isolated by subcellular compartmentalization and/or the association with distinct Dicer complexes. The degree of terminal modification may play an important role in regulating the extent to which primary smRNA signals can be amplified by RNA-dependent RNA polymerases.
Ebhardt
H Alexander
HA
Department of Molecular Biology and Biochemistry, Simon Fraser University, 8888 University Drive, Burnaby, BC, Canada V5A 1S6.
Thi
Emily P
EP
Wang
Ming-Bo
MB
Unrau
Peter J
PJ
D018506:Gene Expression Regulation, Plant / Q000502:physiology*; D008745:Methylation; D035683:MicroRNAs / Q000737:chemistry* / Q000378:metabolism; D010447:Peptide Hydrolases / Q000737:chemistry / Q000378:metabolism; D030821:Plants, Genetically Modified; D034622:RNA Interference / Q000502:physiology*; D018749:RNA, Plant / Q000737:chemistry* / Q000378:metabolism; D034741:RNA, Small Interfering / Q000737:chemistry* / Q000378:metabolism; D012266:Ribose / Q000737:chemistry / Q000378:metabolism; D014026:Nicotiana / Q000737:chemistry* / Q000502:physiology
D035683:MicroRNAs; D018749:RNA, Plant; D034741:RNA, Small Interfering; D012266:Ribose; D010447:Peptide Hydrolases