MicroRNAs (miRNAs) are small, endogenous RNAs that regulate gene expression in plants and animals. In plants, these approximately 21-nucleotide RNAs are processed from stem-loop regions of long primary transcripts by a Dicer-like enzyme and are loaded into silencing complexes, where they generally direct cleavage of complementary mRNAs. Although plant miRNAs have some conserved functions extending beyond development, the importance of miRNA-directed gene regulation during plant development is now particularly clear. Identified in plants less than four years ago, miRNAs are already known to play numerous crucial roles at each major stage of development-typically at the cores of gene regulatory networks, targeting genes that are themselves regulators, such as those encoding transcription factors and F-box proteins.

/pdf/micrornas-and-their-regulatory-roles-in-plants-11zqu0d6p0.pdf

MicroRNAs AND THEIR REGULATORY ROLES IN PLANTS

https://www.ndsu.edu/pubweb/~mcclean/plsc731/homework/khraiwesh%20et%20al%20-%20role%20of%20miRNAs%20and%20siRNAs%20in%20biotic%20and%20abiotic%20stress%20responses%20of%20plants.pdf

Role of miRNAs and siRNAs in biotic and abiotic stress responses of plants

Regulatory small RNAs, which range in size from 20 to 24 nucleotides, are ubiquitous components of endogenous plant transcriptomes, as well as common responses to exogenous viral infections and introduced double-stranded RNA (dsRNA). Endogenous small RNAs derive from the processing of helical RNA precursors and can be categorized into several groups based on differences in biogenesis and function. A major distinction can be observed between small RNAs derived from singlestranded precursors with a hairpin structure [referred to here as hairpin RNAs (hpRNAs)] and those derived from dsRNA precursors [small interfering RNAs (siRNAs)]. hpRNAs in plants can be divided into two secondary groups: microRNAs and those that are not microRNAs. The currently known siRNAs fall mostly into one of three secondary groups: heterochromatic siRNAs, secondary siRNAs, and natural antisense transcript siRNAs. Tertiary subdivisions can be identified within many of the secondary classifications as well. Comparisons between the different classes of plant small RNAs help to illuminate key goals for future research.

/pdf/classification-and-comparison-of-small-rnas-from-plants-456g2wbhbb.pdf

Classification and Comparison of Small RNAs from Plants

Plant genomes encode various small RNAs that function in distinct, yet overlapping, genetic and epigenetic silencing pathways. However, the abundance and diversity of small-RNA classes varies among plant species, suggesting coevolution between environmental adaptations and gene-silencing mechanisms. Biogenesis of small RNAs in plants is well understood, but we are just beginning to uncover their intricate regulation and activity. Here, we discuss the biogenesis of plant small RNAs, such as microRNAs, secondary siRNAs and heterochromatic siRNAs, and their diverse cellular and developmental functions, including in reproductive transitions, genomic imprinting and paramutation. We also discuss the diversification of small-RNA-directed silencing pathways through the expansion of RNA-dependent RNA polymerases, DICER proteins and ARGONAUTE proteins.

/pdf/the-expanding-world-of-small-rnas-in-plants-1ouxzc3mtq.pdf

The expanding world of small RNAs in plants

MicroRNAs (miRNAs) represent an important class of small non-coding RNAs (sRNAs) that regulate gene expression by targeting messenger RNAs. However, assigning miRNAs to their regulatory target genes remains technically challenging. Recently, high-throughput CLIP-Seq and degradome sequencing (Degradome-Seq) methods have been applied to identify the sites of Argonaute interaction and miRNA cleavage sites, respectively. In this study, we introduce a novel database, starBase (sRNA target Base), which we have developed to facilitate the comprehensive exploration of miRNA–target interaction maps from CLIP-Seq and Degradome-Seq data. The current version includes high-throughput sequencing data generated from 21 CLIP-Seq and 10 DegradomeSeq experiments from six organisms. By analyzing millions of mapped CLIP-Seq and Degradome-Seq reads, we identified � 1 million Ago-binding clusters and � 2 million cleaved target clusters in animals and plants, respectively. Analyses of these clusters, and of target sites predicted by 6 miRNA target prediction programs, resulted in our identification of approximately 400 000 and approximately 66 000 miRNA-target regulatory relationships from CLIP-Seq and Degradome-Seq data, respectively. Furthermore, two web servers were provided to discover novel miRNA target sites from CLIP-Seq and Degradome-Seq data. Our web implementation supports diverse query types and exploration of common targets, gene ontologies and pathways. The starBase is available at http://starbase.sysu .edu.cn/.

/pdf/starbase-a-database-for-exploring-microrna-mrna-interaction-3efa3k1418.pdf

starBase: a database for exploring microRNA–mRNA interaction maps from Argonaute CLIP-Seq and Degradome-Seq data

Higher plants have evolved multiple proteins in the RNase III family to produce and regulate different classes of small RNAs with specialized molecular functions. In rice (Oryza sativa), numerous genomic clusters are targeted by one of two microRNAs (miRNAs), miR2118 and miR2275, to produce secondary small interfering RNAs (siRNAs) of either 21 or 24 nucleotides in a phased manner. The biogenesis requirements or the functions of the phased small RNAs are completely unknown. Here we examine the rice Dicer-Like (DCL) family, including OsDCL1, -3a, -3b and -4. By deep sequencing of small RNAs from different tissues of the wild type and osdcl4-1, we revealed that the processing of 21-nucleotide siRNAs, including trans-acting siRNAs (tasiRNA) and over 1000 phased small RNA loci, was largely dependent on OsDCL4. Surprisingly, the processing of 24-nucleotide phased small RNA requires the DCL3 homolog OsDCL3b rather than OsDCL3a, suggesting functional divergence within DCL3 family. RNA ligase-mediated 5' rapid amplification of cDNA ends and parallel analysis of RNA ends (PARE)/degradome analysis confirmed that most of the 21- and 24-nucleotide phased small RNA clusters were initiated from the target sites of miR2118 and miR2275, respectively. Furthermore, the accumulation of the two triggering miRNAs requires OsDCL1 activity. Finally, we show that phased small RNAs are preferentially produced in the male reproductive organs and are likely to be conserved in monocots. Our results revealed significant roles of OsDCL4, OsDCL3b and OsDCL1 in the 21- and 24-nucleotide phased small RNA biogenesis pathway in rice.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/j.1365-313X.2011.04805.x

Roles of DCL4 and DCL3b in rice phased small RNA biogenesis

MicroRNAs (miRNAs) and small interfering RNAs (siRNAs) are two types of noncoding RNAs involved in developmental regulation, genome maintenance, and defense in eukaryotes. The activity of Dicer or Dicer-like (DCL) proteins is required for the maturation of miRNAs and siRNAs. In this study, we cloned and sequenced 66 candidate rice (Oryza sativa) miRNAs out of 1,650 small RNA sequences (19 to approximately 25 nt), and they could be further grouped into 21 families, 12 of which are newly identified and three of which, OsmiR528, OsmiR529, and OsmiR530, have been confirmed by northern blot. To study the function of rice DCL proteins (OsDCLs) in the biogenesis of miRNAs and siRNAs, we searched genome databases and identified four OsDCLs. An RNA interference approach was applied to knock down two OsDCLs, OsDCL1 and OsDCL4, respectively. Strong loss of function of OsDCL1IR transformants that expressed inverted repeats of OsDCL1 resulted in developmental arrest at the seedling stage, and weak loss of function of OsDCL1IR transformants caused pleiotropic developmental defects. Moreover, all miRNAs tested were greatly reduced in OsDCL1IR but not OsDCL4IR transformants, indicating that OsDCL1 plays a critical role in miRNA processing in rice. In contrast, the production of siRNA from transgenic inverted repeats and endogenous CentO regions were not affected in either OsDCL1IR or OsDCL4IR transformants, suggesting that the production of miRNAs and siRNAs is via distinct OsDCLs.

Loss of Function of OsDCL1 Affects MicroRNA Accumulation and Causes Developmental Defects in Rice

MicroRNAs (miRNAs) and small interfering RNAs (siRNAs) are two types of noncoding RNAs involved in developmental regulation, genome maintenance, and defense in eukaryotes The activity of Dicer or Dicer-like (DCL) proteins is required for the maturation of miRNAs and siRNAs In this study, we cloned and sequenced 66 candidate rice (Oryza sativa) miRNAs out of 1,650 small RNA sequences (19 to approximately 25 nt), and they could be further grouped into 21 families, 12 of which are newly identified and three of which, OsmiR528, OsmiR529, and OsmiR530, have been confirmed by northern blot To study the function of rice DCL proteins (OsDCLs) in the biogenesis of miRNAs and siRNAs, we searched genome databases and identified four OsDCLs An RNA interference approach was applied to knock down two OsDCLs, OsDCL1 and OsDCL4, respectively Strong loss of function of OsDCL1IR transformants that expressed inverted repeats of OsDCL1 resulted in developmental arrest at the seedling stage, and weak loss of function of OsDCL1IR transformants caused pleiotropic developmental defects Moreover, all miRNAs tested were greatly reduced in OsDCL1IR but not OsDCL4IR transformants, indicating that OsDCL1 plays a critical role in miRNA processing in rice In contrast, the production of siRNA from transgenic inverted repeats and endogenous CentO regions were not affected in either OsDCL1IR or OsDCL4IR transformants, suggesting that the production of miRNAs and siRNAs is via distinct OsDCLs Most eukaryotes have two classes of short (21‐25 nt) noncoding RNAs, microRNAs (miRNAs) and small interfering RNAs (siRNAs), which are involved in RNAsilencing pathways (Baulcombe, 2004) The two classes of small RNAs are related but differ in their biogenesis from their origins, initiation, to their subsequent assembly into different RNA-induced silencing complexes (RISCs; Tang, 2005) miRNAs are processed from single-stranded RNAs with imperfect stem-loop structure, whereas siRNAs are derived from doublestranded RNAs (dsRNAs) Both miRNA and siRNA precursors are cut into double-stranded duplexes by Dicer, a multidomian enzyme of the RNase III family Then the duplexes are unwound and one strand is subsequently assembled into miRISC or siRISC, respectively Both miRNAs and siRNAs down-regulate gene expression via sequence complementarity to their target mRNAs by either cleavage or translational re

Loss of Function of OsDCL1 Affects MicroRNA Accumulation and Causes Developmental

MicroRNAs (miRNAs) and small interfering RNAs (siRNAs) regulate gene expression in eukaryotes. Plant miRNAs modulate their targets mainly via messenger RNA (mRNA) cleavage. Small RNA (sRNA) targets have been extensively investigated in Arabidopsis using computational prediction, experimental validation, and degradome sequencing. However, small RNA targets are largely unknown in rice (Oryza sativa). Here, we report global identification of small RNA targets using high throughput degradome sequencing in the rice indica cultivar 93–11 (Oryza sativa L. ssp. indica). One hundred and seventy-seven transcripts targeted by a total of 87 unique miRNAs were identified. Of targets for the conserved miRNAs between Arabidopsis and rice, transcription factors comprise around 70% (58 in 82), indicating that these miRNAs act as masters of gene regulatory nodes in rice. In contrast, non-conserved miRNAs targeted diverse genes which provide more complex regulatory networks. In addition, 5 AUXIN RESPONSE FACTORs (ARFs) cleaved by the TAS3 derived ta-siRNAs were also detected. A total of 40 sRNA targets were further validated via RNA ligasemediated 5’ rapid amplification of cDNA ends (RLM 5’-RACE). Our degradome results present a detailed sRNA-target interaction atlas, which provides a guide for the study of the roles of sRNAs and their targets in rice.

Pingchuan Li

Papers

Roles of DCL4 and DCL3b in rice phased small RNA biogenesis

Loss of Function of OsDCL1 Affects MicroRNA Accumulation and Causes Developmental Defects in Rice

Loss of Function of OsDCL1 Affects MicroRNA Accumulation and Causes Developmental

Degradome sequencing reveals endogenous small RNA targets in rice ( Oryza sativa L. ssp. indica )

Identification and expression analysis of miRNAs from nitrogen-fixing soybean nodules