Novel microRNA-target RNA pairs revealed by Parallel analysis of RNA ends. Arabidopsis thaliana

MicroRNAs are important regulatory molecules in most eukaryotes and the identification of their mRNA targets is essential for their functional analysis. From inflorescence tissue of Arabidopsis, >28,000,000 signatures were sequenced from 5’ ends of polyadenylated products of mRNA decay. Within the set of ~27,000 transcripts included in the 3,500,000 non-redundant signatures, several previously predicted but non-validated miRNA targets were found. Like validated targets, most showed a single abundant signature at the miRNA cleavage site, particularly in libraries from a mutant deficient in the 5’ to 3’ exonuclease AtXRN4. Among the most unexpected miRNA targets discovered were miRNA precursor transcripts that are self-targeted by their own mature miRNA. Although the miRNAs of Arabidopsis have been extensively investigated, working in reverse from the cleaved targets, additional novel miRNAs were identified and validated. This deep and versatile approach will impact the study of other aspects of RNA processing beyond miRNA-target RNA pair analyses. Keywords: miRNA-target RNA pairs, Palallel analysis of RNA ends, PARE, SBS Overall design: RNA from inflorescence tissue from wildtype Arabidopsis and xrn4 mutants was extracted. Following polyA RNA extraction, libraries for PARE were constructed. RNA from inflorescence tissue from wildtype Arabidopsis (Col-0), and rdr2 and dcl2,3,4 mutants was extracted. The material was submitted to Illumina and libraries were constructed for small RNAs. Individual small RNA libraries from inflorescences of Arabidopsis wildtype, rdr2, and dcl2/dcl3/dcl4 were sequenced with SBS at Illumina, Inc. The distribution of small RNA sizes in each library is in agreement with that published previously for the wild type or the respective mutants sequenced with MPSS or 454; however, the presence and abundances of some small RNAs may differ. This may be attributed to the use of different sequencing technologies with different depths, biological differences in the tissue used, or other unknown reasons. raw data requested but not provided for GSM280226 and GSM280227