Identification of browning-related microRNAs and their targets reveals complex miRNA-mediated browning regulatory networks in Luffa cylindrica

As a non-coding and endogenous small RNA, MicroRNA (miRNA) takes a vital regulatory role in plant growth and development. Long-term storage and processing of many fruits and vegetables, including Luffa, are subject to influences from browning, a common post-harvest problem that adversely affects flavor, aroma, and nutritional value. The browning regulatory networks mediated by miRNA, however, remain largely unexplored. For a systematic identification of browning-responsive miRNAs and the targets, we built two RNA libraries from Luffa pulps of near-isogenic line, with resistant and sensitive browning characteristics respectively, and then sequenced them using Solexa high-throughput technology. We consequently identified 179 known miRNAs that represent 17 non-conserved miRNA families and 24 conserved families, as well as 84 potential novel miRNAs, among which 16 miRNAs (eight known and eight novel miRNAs) were found to exhibit significant differential expressions and were thus identified as browning-related miRNAs. We then studied those browning-responsive miRNAs and the corresponding targets with RT-qPCR and finally validated their expression patterns. The results revealed that the expression patterns are specific to plant development stages and the miRNAs are identified with 39 target transcripts, which involve in plant development, defense response, transcriptional regulation, and signal transduction. After characterizing these miRNAs and their targets, we propose a browning regulatory network model of miRNA-mediatation in this paper. The findings of the work are helpful for the understanding of miRNA-mediated regulatory mechanisms of browning in Luffa, and will facilitate genetic improvement of pulp characteristics in Luffa.