A non-canonical plant microRNA target site

Abstract

Plant microRNAs (miRNAs) typically form near-perfect duplexes with their targets and mediate mRNA cleavage. Here, we describe an unconventional miRNA target of miR398 in Arabidopsis, an mRNA encoding the blue copper-binding protein (BCBP). BCBP mRNA carries an miR398 complementary site in its 5′-untranslated region (UTR) with a bulge of six nucleotides opposite to the 5′ region of the miRNA. Despite the disruption of a target site region thought to be especially critical for function, BCBP mRNAs are cleaved by ARGONAUTE1 between nucleotides 10th and 11th, opposite to the miRNA, like conventional plant target sites. Levels of BCBP mRNAs are inversely correlated to levels of miR398 in mutants lacking the miRNA, or transgenic plants overexpressing it. Introducing two mutations that disrupt the miRNA complementarity around the cleavage site renders the target cleavage-resistant. The BCBP site functions outside of the context of the BCBP mRNA and does not depend on 5′-UTR location. Reducing the bulge does not interfere with miR398-mediated regulation and completely removing it increases the efficiency of the slicing. Analysis of degradome data and target predictions revealed that the miR398-BCBP interaction seems to be rather unique. Nevertheless, our results imply that functional target sites with non-perfect pairings in the 5′ region of an ancient conserved miRNA exist in plants.

Revisiting Criteria for Plant MicroRNA Annotation in the Era of Big Data

“…In the Brassicaceae and Cleomaceae (both in the Rosids supergroup of eudictos), miR396 also targets Basic homologs (Debernardi et al, 2012), while in sunflower (Helianthus annus, in the asterid supergroup of eudicots) it has gained a WRKY transcription factor target (Giacomelli et al, 2012). Gains of lineage-specific targets have also been described for the ancient miR156, miR159, miR167, miR398, miR408, and miR482 families Buxdorf et al, 2010;Chorostecki et al, 2012;Brousse et al, 2014;Xia et al, 2015b). These lineage-specific targets frequently have complementarity patterns that include bulged nucleotides, rendering them undetectable by standard plant miRNA target identification software.…”

“…Alternative approaches that are more sensitive to these diverse complementarity sites have been described to meet this challenge (Chorostecki et al, 2012;Brousse et al, 2014).…”

Literature review of baseline information to support the risk assessment of RNAi‐based GM plants

“…Central mismatches interfere with repression . However, miR398 in Arabidopsis binds 5'UTR of the blue copper-binding protein mRNA with a bulge of six nucleotides opposite to the 5' region of the miRNA (Brousse et al, 2014). These and other studies led to consensus base pairing rules for a functional plant miRNA-target interaction: little tolerance for mismatches at positions 2-13, with especially little tolerance of mismatches at positions 9-11, and more tolerance of mismatches at positions 1, and 14-21 (Wang et al, 2015a).…”

Genome-wide identification of turnip mosaic virus-responsive microRNAs in non-heading Chinese cabbage by high-throughput sequencing

“…php?GDB = Pe) as the "Section 2" described. Previous study had found that besides the opening reading frame, miRNAs can also target the 5-UTR or 3-UTR domain of some genes (Brousse et al, 2014), thus these types of candidate target genes were included in our study. Then a total of putative 271 target genes were obtained for the identified conserved and novel miRNAs (Supplementary Tables S7 and S8).…”