Identification of microRNA targets in tomato fruit development using high-throughput sequencing and degradome analysis.
ABSTRACT MicroRNAs (miRNAs) play important roles in plant development through regulation of gene expression by mRNA degradation or translational inhibition. Despite the fact that tomato (Solanum lycopersicum) is the model system for studying fleshy fruit development and ripening, only a few experimentally proven miRNA targets are known, and the role of miRNA action in these processes remains largely unknown. Here, by using parallel analysis of RNA ends (PARE) for global identification of miRNA targets and comparing four different stages of tomato fruit development, a total of 119 target genes of miRNAs were identified. Of these, 106 appeared to be new targets. A large part of the identified targets (56) coded for transcription factors. Auxin response factors, as well as two known ripening regulators, COLORLESS NON-RIPENING (CNR) and APETALA2a (SlAP2a), with developmentally regulated degradation patterns were identified. The levels of the intact messenger of both CNR and AP2a are actively modulated during ripening, by miR156/157 and miR172, respectively. Additionally, two TAS3-mRNA loci were identified as targets of miR390. Other targets such as ARGONAUTE 1 (AGO1), shown to be involved in miRNA biogenesis in other plant species, were identified, which suggests a feedback loop regulation of this process. In this study, it is shown that miRNA-guided cleavage of mRNAs is likely to play an important role in tomato fruit development and ripening.
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ABSTRACT: Background MicroRNAs (miRNAs) are small (approximately 21 nucleotide) non-coding RNAs that are key post-transcriptional gene regulators in eukaryotic organisms. More than 100 cassava miRNAs have been identified in a conservation analysis and a repertoire of cassava miRNAs have also been characterised by next-generation sequencing (NGS) in recent studies. Here, using NGS, we profiled small non-coding RNAs and mRNA genes in two cassava cultivars and their wild progenitor to identify and characterise miRNAs that are potentially involved in plant growth and starch biosynthesis.ResultsSix small RNA and six mRNA libraries from leaves and roots of the two cultivars, KU50 and Arg7, and their wild progenitor, W14, were subjected to NGS. Analysis of the sequencing data revealed 29 conserved miRNA families and 33 new miRNA families. Together, these miRNAs potentially targeted a total of 360 putative target genes. Whereas 16 miRNA families were highly expressed in cultivar leaves, another 13 miRNA families were highly expressed in storage roots of cultivars. Co-expression analysis revealed that the expression level of some targets had negative relationship with their corresponding miRNAs in storage roots and leaves; these targets included MYB33, ARF10, GRF1, RD19, APL2, NF-YA3 and SPL2, which are known to be involved in plant development, starch biosynthesis and response to environmental stimuli.Conclusion The identified miRNAs, target mRNAs and target gene ontology annotation all shed light on the possible functions of miRNAs in Manihot species. The differential expression of miRNAs between cultivars and their wild progenitor, together with our analysis of GO annotation and confirmation of miRNA:target pairs, might provide insight into know the differences between wild progenitor and cultivated cassava.BMC Plant Biology 02/2015; 15(1):33. DOI:10.1186/s12870-014-0355-7 · 3.94 Impact Factor
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ABSTRACT: MicroRNAs (miRNAs) are a class of small endogenous non-coding RNAs that modulate gene expression in both plants and animals and are involved in several biological processes, ranging from organ differentiation to biotic and abiotic stress responses. We identified two cold stress response microRNAs that showed differential expression in Solanum lycopersicum plants subjected to cold stress. We observed that Sly-miR166 and Sly-miR319 were up-regulated by cold treatments. The up-regulation of Sly-miR166 and Sly-miR319 in cold stress-treated S. lycopersicum seedlings and the down-regulation of their respective targets, HD-Zip III and GAMyb-like that validate by 5′-RACE technique, suggests that these miRNAs play a critical role in regulating S. lycopersicum responses to cold stress.Plant Biotechnology Reports 07/2014; 8(4):349-356. DOI:10.1007/s11816-014-0326-3 · 1.59 Impact Factor
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ABSTRACT: In plants, microRNAs (miRNAs) play essential roles in growth, development, yield, stress response and interactions with pathogens. However no miRNA has been experimentally documented to be functionally involved in fruit ripening although many miRNAs have been profiled in fruits. Here we show that SlymiR157 and SlymiR156 differentially modulate ripening and softening in tomato (Solanum lycopersicum). SlymiR157 is expressed and developmentally regulated in normal tomato fruits and in those of the Colourless non-ripening (Cnr) epimutant. It regulates expression of the key ripening gene LeSPL-CNR in a likely dose-dependent manner through miRNA-induced mRNA degradation and translation repression. Viral delivery of either pre-SlymiR157 or mature SlymiR157 results in delayed ripening. Furthermore, qRT-PCR profiling of key ripening regulatory genes indicates that the SlymiR157-target LeSPL-CNR may affect expression of LeMADS-RIN, LeHB1, SlAP2a and SlTAGL1. However SlymiR156 does not affect the onset of ripening, but it impacts fruit softening after the red ripe stage. Our findings reveal that working together with a ripening network of transcription factors, SlymiR157 and SlymiR156 form a critical additional layer of regulatory control over the fruit ripening process in tomato.Scientific Reports 01/2015; 5. DOI:10.1038/srep07852 · 5.08 Impact Factor