Introduction
According to the central dogma of biology, DNAs encode the genetic information required to make proteins, and RNAs are messenger molecules that ensure the genetic information is translated into proteins on ribosomes. Although this classic model has been a major principle of molecular biology for more than half a century, findings about noncoding RNAs in the past few decades have shed light on the essential functions exerted by genomic regions previously thought of as “junk” DNA [1–5]. Small RNAs are a group of 20–30 nucleotide (nt) noncoding RNAs that exist in diverse eukaryotic organisms [1–3]. Despite their tiny size, small RNAs affect numerous biological processes throughout the lives of living organisms by transcriptionally or post-transcriptionally regulating gene expression [6–9]. Due to the widespread impact of small RNAs and their functional potentials as powerful molecular biology tools, tremendous research efforts have focused on the small RNA regulatory machinery, small RNA metabolism, and the underlying mechanisms. Plenty of recent studies have revealed the functions of small RNAs in diseases such as cancer, further underscoring the urgency and importance of deciphering small RNA-related mechanisms [10–12].
Discovery history of small RNAs
Although the essential functions of small RNAs in gene expression regulation and the mechanisms underlying small RNA metabolism were not revealed until two decades ago, the phenomena resulting from regulation mediated by endogenous and transgene-derived small RNAs were already reported in numerous studies in the late 1980s.
The first report of an endogenous small RNA was in 1993; the microRNA (miRNA) lin-4 was identified in the nematode Caenorhabditis elegans (C. elegans) through molecular genetic analysis showing that a mutation in the miRNA gene led to developmental defects [13]. In the same year, the first small RNA/target module, lin4/lin14, was also identified [14]. A novel mechanism of gene regulation at the post-transcriptional level was established by these two landmark discoveries. However, the broader importance of miRNAs was not fully recognized until 7 years later, when let-7, a 21-nt C. elegans miRNA, was found to regulate the L4-to-adult developmental transition in larva [15]. Shortly thereafter, a large class of small regulatory RNAs exhibiting diverse expression patterns was uncovered in nematodes [16–18].
In plants, the phenomenon of co-suppression was discovered in studies of flavanol and anthocyanin biosynthesis. In the petunia, an overexpression of a genes encoding dihydroflavonol 4-reductase and chalcone synthase led to reduced RNA levels of both the endogenous genes and the transgenes [19–21]. A few years later in 1999, a class of short antisense small interfering RNA (siRNA) was discovered and characterized as the specific determinant of post-transcriptional gene silencing (PTGS) [22]. In 2002, a study using a small RNA cloning strategy, miRNAs in Arabidopsis and rice were found [23], and their corresponding target mRNAs with near-perfect sequence complementarity were predicted [24].
As with the findings in nematodes, the observed silencing effects in plants were similarly mediated by small noncoding RNAs, and these processes were found to be regulated by similar enzymes in most eukaryotes [25–27]. Since the initial discoveries, numerous small RNAs have been identified in diverse eukaryotes through high-throughput sequencing and bioinformatic prediction strategies [28–30].
Diversity of small RNAs
Small RNAs are classified into three major types based on their biogenesis and associated proteins: miRNAs, siRNAs, and Piwi-interacting RNAs (piRNAs), which only present in animals. miRNAs are generated from stem-loop- or hairpin-structured single-stranded RNA (ssRNA) precursors. The processing of precursors into mature miRNAs requires the activity of RNase III-type endonucleases DICER and DROSHA in animals or DICER-LIKE (DCL) in plants [27, 31, 32]. The biogenesis of siRNAs also requires DICERs or DCLs, but siRNAs derive from RNA-DEPENDENT RNA POLYMERASEs (RDRs)-produced long double-stranded RNA (dsRNA) precursors with perfect complementarity or single-stranded transcripts with inverted repeat elements [33]. siRNAs can be further divided into several subgroups: heterochromatic small interfering RNA (hc-siRNA), natural antisense transcript-derived small interfering RNAs (nat-siRNA), and phased small interfering RNAs (phasiRNAs). Despite their different origins, both miRNAs and siRNAs are loaded into ARGONAUTE (AGO) proteins to induce gene silencing.
In terms of their mode of action, small RNAs specifically recognize target transcripts through sequence complementarity and mediate transcriptional gene silencing (TGS) or PTGS. In TGS, small RNAs direct DNA methylation and histone modification to inhibit the transcription of transgenes or transposons, thereby contributing to genome stability maintenance [33]. In PTGS, small RNAs modulate gene expression via transcript cleavage, decay, or translation inhibition of target RNAs that derive from protein-coding genes, transposable elements, transgenes, and viral RNAs [2, 8, 9]. Here we discuss the biogenesis, movements, regulatory functions, and action modes of major small RNA groups involved in PTGS in plants.
miRNAs
In plants, miRNAs and siRNAs are the two major classes of small RNAs [2]. Despite that miRNAs constitute a tiny fr...