Dicer-like 1 (DCL1) is a core component of the plant microRNA (miRNA) biogenesis machinery. MiRNA is transcribed as a precursor RNA, termed primary miRNA (pri-miRNA), which is cleaved by DCL1 in two steps to generate miRNA/miRNA* duplex. Pri-miRNA is a single-stranded RNA that forms a hairpin structure with a number of unpaired bases, hereafter called mismatches, on its stem. In the present study, by using purified recombinant Arabidopsis DCL1, we presented evidence that mismatches on the stem of pri-miRNA are important for precise DCL1 cleavage. We showed that a mismatch at the loop-distal side of the end of miRNA/miRNA* duplex is important for efficient cleavage of pri-miRNA in vitro, as previously suggested in planta. On the contrary, mismatches distant from the miRNA/miRNA* duplex region are important for determining the cleavage position by DCL1. The purified DCL1 proteins cleaved mutant pri-miRNA variants without such mismatches at a position at which wild-type pri-miRNA variants are not usually cleaved, resulting in an increased accumulation of small RNA different from miRNA. Therefore, our results suggest that, in addition to the distance from the ssRNA-dsRNA junction, mismatches on the stem of pri-miRNA function as a determinant for precise processing of pri-miRNA by DCL1 in plants.
Bibliographical noteFunding Information:
This work was supported by JSPS KAKENHI Grand Numbers JP17K15416 to Y.I. and JP20J15594 to R.H.
This work was supported by JSPS KAKENHI Grand Numbers JP17K15416 to Y.I. and JP20J15594 to R.H. The authors thank The Arabidopsis Information Resource (TAIR) for the gene annotation data. We thank Ayumi Sakei (Osaka Prefecture University) for technical assistance.
© 2022 Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd.
- Arabidopsis thaliana
- microRNA biogenesis
- RNA secondary structure
ASJC Scopus subject areas
- Cell Biology