Genome-Wide Analysis Reveals Diversity of Rice Intronic miRNAs in Sequence Structure, Biogenesis and Function

Yong-Ao Tong,Linhong Fan,Shenghua Wang,Cheng Zhan,Taobo Ai,Hua Peng,Ying Xu

doi:10.1371/journal.pone.0063938

Abstract

Intronic microRNAs (in-miRNAs) as a class of miRNA family that regulates gene expression are still poorly understood in plants. In this study, we systematically identified rice in-miRNAs by re-mining eight published small RNA-sequencing datasets of rice. Furthermore, based on the collected expression, annotation, and putative target data, we investigated the structures, potential functions, and expression features of these in-miRNAs and the expression patterns of their host genes. A total of 153 in-miRNAs, which account for over 1/4 of the total rice miRNAs, were identified. In silico expression analysis showed that most of them (∼63%) are tissue or stage-specific. However, a majority of their host genes, especially those containing clustered in-miRNAs, exhibit stable high-level expressions among 513 microarray datasets. Although in-miRNAs show diversity in function and mechanism, the DNA methylation directed by 24 nt in-miRNAs may be the main pathway that controls the expressions of target genes, host genes, and even themselves. These findings may enhance our understanding on special functions of in-miRNAs, especially in mediating DNA methylation that was concluded to affect the stability of expression and structure of host and target genes.

Highlights

MicroRNAs are,22 nt endogenous small RNAs that have gained the worldwide attention from researchers in gene expression regulation and gene therapy because of their vital role in the regulation of development, metabolism, stress responses, and pathogen responses of unicellular [1] and multicellular organisms [2]
Novel miRNAs in Intronic Regions of the Rice Genome The strategy used in this paper in identifying miRNAs in the intronic regions of the rice genome is as follows: (a) The inverted repeat (IR) were picked up from the intronic sequences of the rice genome downloaded from the MSU Rice Genome Annotation Project database. (b) The IRs with low-abundant expressions were filtered out by checking their total reads from all eight sequencing samples. (c) The secondary structures of IRs were assessed using RNAFold [31]. (d) Mature miRNA and miRNA* were extracted from pre-miRNA candidates using our custom script
We have identified 40 novel miRNAs derived from 20 intronic pre-miRNAs and 34 novel miRNAs from 18 annotated pre-miRNAs from the intronic regions of protein-coding genes of the rice genome using a comprehensive analysis of eight sets of small RNA sequencing data from the NCBI and gene annotation data from the MSU Rice Genome Annotation Project (Release 6.0; http://rice.plantbiologu.msu.edu) [28]

Summary

Introduction

MicroRNAs (miRNAs) are ,22 nt endogenous small RNAs that have gained the worldwide attention from researchers in gene expression regulation and gene therapy because of their vital role in the regulation of development, metabolism, stress responses, and pathogen responses of unicellular [1] and multicellular organisms [2]. We identified 74 novel miRNAs from eight published small RNA-sequencing datasets by systematical investigation of intronic regions of protein coding genes in the rice genome.

Results

Conclusion