Identification of conserved and novel miRNAs responsive to heat stress in flowering Chinese cabbage using high-throughput sequencing

Plant microRNAs (miRNAs) are noncoding and endogenous key regulators that play significant functions in regulating plant responses to stress, and plant growth and development. Heat stress is a critical abiotic stress that reduces the yield and quality of flowering Chinese cabbage (Brassica campestris L. ssp. chinensis var. utilis Tsen et Lee). However, limited information is available on whether miRNAs are involved in the regulation of heat stress in B. campestris. A high-throughput sequencing approach was used to identify novel and conserved heat-responsive miRNAs in four small RNA libraries of flowering Chinese cabbage using leaves collected at 0 h, 1 h, 6 h and 12 h after a 38 °C heat-stress treatment. The analysis identified 41 conserved miRNAs (belonging to 19 MIR families), of which MIR156, MIR159, MIR168, MIR171 and MIR1885 had the most abundant molecules. Prediction and evaluation of novel miRNAs using the unannotated reads resulted in 18 candidate miRNAs. Differential expression analysis showed that most of the identified miRNAs were downregulated in heat-treated groups. To better understand functional importance, bioinformatic analysis predicted 432 unique putative target miRNAs involved in cells, cell parts, catalytic activity, cellular processes and abiotic stress responses. Furthermore, the Kyoto Encyclopedia of Genes and Genomes maps of flowering Chinese cabbage identified the significant role of miRNAs in stress adaptation and stress tolerance, and in several mitogen-activated protein kinases signaling pathways including cell death. This work presents a comprehensive study of the miRNAs for understanding the regulatory mechanisms and their participation in the heat stress of flowering Chinese cabbage.