Background MicroRNAs (miRNAs) are endogenous single-stranded small RNAs that regulate the

Background MicroRNAs (miRNAs) are endogenous single-stranded small RNAs that regulate the expression of specific mRNAs involved in diverse biological processes. Ath-miR774, led to the DCL1-dependent accumulation of both miRNAs and down-regulation of their different mRNA targets encoding F-box proteins. Conclusions In addition to polycistronic precursors carrying related miRNAs, plants also contain precursors allowing coordinated expression of non-homologous miRNAs to co-regulate functionally related target transcripts. This mechanism paves the way for using polycistronic MIRNA precursors as a new molecular tool for plant biologists to simultaneously control the expression of different genes. Background MicroRNAs (miRNAs) are endogenous approximately 21-nucleotide single-stranded small RNAs derived from MIRNA precursors that are able to fold-back into a stable secondary structure (stem loop Mouse monoclonal to CEA or hairpin). 935881-37-1 manufacture miRNAs act in many developmental processes as well as environmental and pathogenic responses [1-4] through the post-transcriptional regulation of target mRNAs. These targets carry a sequence-specific miRNA recognition site, leading to transcript cleavage and/or inhibition of mRNA translation [1,5,6]. Primary miRNA transcripts (pri-MIRNA) are transcribed by RNA polymerase II, and several ribonucleoprotein (RNP) complexes are involved in their maturation, a process that differs between animals and plants [1,6-11]. In animals, formation of an approximately 21-bp miRNA-miRNA* duplex successively involves two RNase III enzymatic complexes: the Drosha enzyme, which cleaves long pri-MIRNA in the nucleus to generate short (approximately 70- to 80-nucleotide) hairpins (so called pre-MIRNA) and the Dicer enzyme, which produces the miRNA after cytoplasmic export of pre-MIRNAs 935881-37-1 manufacture through Exportin 5 [11]. In plants, however, both cleavages are likely nuclear localized and involve a single Dicer-like enzyme 1 (DCL1) complex [6,9,10]. The miRNA-miRNA* duplex is exported to the cytoplasm by HASTY, the plant ortholog of Exportin 5 [12,13]. Subsequently, these duplexes are converted into single-stranded miRNAs upon incorporation into an ARGONAUTE (AGO) ribonucleoprotein complex, referred to as the RNA-induced silencing complex (RISC). The miRNAs guide sequence-specific cleavage and/or translational repression of target transcripts into the RISC complex [6,9-11]. Recent deep sequencing of plant small RNA libraries has led to the identification of more than 1,300 miRNAs in various plants (miRBase, release 13.0, March 2009) [14]. Based on comparison of all available plant genomes (even partial ones; 16 genera referenced in miRBase), evolutionarily conserved and non-conserved miRNAs have been proposed. The non-conserved miRNAs have probably emerged in recent evolutionary time scales, and show a wide diversity compared to the restricted number of conserved miRNAs [15]. Indeed, only 5 935881-37-1 manufacture miRNA families are found in more than 40 plant species whereas 25 exist in more than one plant genus [16]. The three higher plant models showing the most comprehensive description of their miRNome are rice (Oryza sativa; 377 MIRNAs), poplar (Populus trichocarpa; 234 MIRNAs) and Arabidopsis (Arabidopsis thaliana; 187 MIRNAs), with 22 families ‘conserved’ between them (indicated in bold in Additional data file 1 based on miRBase 13.0). The numerous non-conserved miRNAs are thus likely to play species-specific roles [15]. Plant and animal MIRNA genes differ in their genomic location and organization. Most plant miRNAs are encoded in intergenic loci, whereas animal miRNAs are also frequently encoded within introns of protein coding genes [17-19]. Plant miRNAs are mainly generated from independent transcriptional units, whereas in Drosophila, nematodes, zebrafish and mammals, around 40 to 50% of the predicted MIRNA genes are located within clusters that are often evolutionarily conserved [18-27]. A maximal distance of 3 kb between two consecutive miRNAs has been used as a stringent criterion to estimate cluster numbers [18]. Clusters in animal genomes usually encode two to three miRNAs but some encode up to eight. Even larger miRNA clusters were predicted in human and zebrafish, containing more than 40 MIRNA loci [18,25,26]. In these clusters, miRNAs are encoded either in independent hairpins or sometimes in both arms of the same hairpin [28]. In plants, even though no systematic analysis of miRNA clusters has been performed in the different available genomes, a few miRNA clusters have been reported [16,29-33]. Clustered miRNAs can be either simultaneously transcribed into a single polycistronic transcript or independently transcribed [1,28,34]. Short distances between consecutive MIRNA loci and coordinated expression of clustered.