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Analysis of Brachypodium miRNA targets: Evidence for diverse control during stress and conservation in bioenergy crops

ABSTRACT: Background: Since the proposal of Brachypodium distachyon as a model for the grasses over 500 Bdi-miRNAs have been annotated in miRBase making Brachypodium second in number only to rice. Other monocots, such as switchgrass, are completely absent from the miRBase database. While a significant number of miRNAs have been identified which are highly conserved across plants, little research has been done with respect to the conservation of miRNA targets. Plant responses to abiotic stresses are regulated by diverse pathways many of which involve miRNAs; however, it can be difficult to identify miRNA guided gene regulation when the miRNA is not the primary regulator of the target mRNA. Results: To investigate miRNA target conservation and stress response involvement, a set of PARE (Parallel Analysis of RNA Ends) libraries totaling over 2 billion reads was constructed and sequenced from Brachypodium, switchgrass, and sorghum representing the first public release of degradome data from the latter two species. Analysis of this data provided not only PARE evidence for miRNA guided cleavage of over 7,000 predicted target mRNAs in Brachypodium, but also evidence for miRNA guided cleavage of over 1,000 homologous transcripts in sorghum and switchgrass. A pipeline was constructed to compare RNA-seq and PARE data made from Brachypodium plants exposed to various abiotic stress conditions. This resulted in the identification of 44 miRNA targets which exhibit stress regulated cleavage. Time course experiments were performed to reveal the relationship between miR393ab, miR169a, miR394ab, and their respective targets throughout the first 36 hours of the cold stress response in Brachypodium. Conclusions: Knowledge gained from this study provides considerable insight into the degradomes and the breadth of miRNA target conservation among these three species. Additionally associations of a number of miRNAs and target mRNAs with the stress responses have been revealed which will aid researchers in developing stress tolerant transgenic crops.

ORGANISM(S): Brachypodium distachyon Panicum virgatum Sorghum bicolor

PROVIDER: GSE112296 | GEO | 2019/01/01

REPOSITORIES: GEO

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Project description:MicroRNAs (miRNAs) are a class of small RNAs which typically function by guiding cleavage of target messenger RNAs. They have been shown to play major roles in a variety of plant processes including development, and responses to pathogens and environmental stresses. To identify new miRNAs and regulation in Arabidopsis thaliana, 27 small RNA libraries were constructed and sequenced from various tissues, stresses and small RNA biogenesis mutants, resulting in 95 million genome-matched sequences. The use of rdr2 to enrich the miRNA population greatly enhanced this analysis and led to the discovery of 44 new miRNAs arising from both known and new precursors. Parallel Analysis of RNA Ends (PARE) data provide evidence that the majority guide target cleavage. The inclusion of novel stress/tissue conditions, such as submergence-stressed flowers, enabled identification of new stress regulation of both miRNAs and their targets, all of which were validated in wild type plants. By combining small RNA expression analysis with ARGONAUTE (AGO) immunoprecipitation data and global target cleavage data from PARE, a much more complete picture of Arabidopsis miRNAs was obtained. This combinatorial approach led to the discovery of AGO loading and target cleavage biases, which gave important insights into tissue-specific expression patterns, pathogen responses and the role of sequence variation among closely related miRNA family members. Examination of various tissues, stresses and small RNA biogenesis mutants in Arabidopsis by high-throughput sequencing for small RNA profiling. We have used AGO-IP and PARE data from the published data, which were downloaded from NCBI GEO with the following accession number. AGO-IP from GSM253622, GSM707682, GSM642335, GSM642336, GSM512703, GSM512702, GSM707683, GSM707684, GSM707685, GSM149080, GSM253623, GSM304283, GSM642337, GSM642338, GSM253624, GSM415788, GSM707686, GSM707687, GSM707688, GSM707689, GSM415787, GSM149081, GSM253625, GSM415789, GSM415790, GSM304285, GSM415791, GSM415792 PARE sequencing data from xrn4 flowers were obtained from Gene Expression Omnibus with accession number GSM280227.

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Analysis of Brachypodium miRNA targets: Evidence for diverse control during stress and conservation in bioenergy crops

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