Identification of miRNAs and target genes regulating catechin biosynthesis in tea (Camellia sinensis)

Abstract

MicroRNAs (miRNAs) are endogenous non-protein-coding small RNAs that play crucial and versatile regulatory roles in plants. Using a computational identification method, we identified 55 conserved miRNAs in tea (Camellia sinensis) by aligning miRNA sequences of different plant species with the transcriptome library of tea strain 1005. We then used quantitative real-time PCR (qRT-PCR) to analyze the expression of 31 identified miRNAs in tea leaves of different ages, thereby verifying the existence of these miRNAs and confirming the reliability of the computational identification method. We predicted which miRNAs were involved in catechin synthesis using psRNAtarget Software based on conserved miRNAs and catechin synthesis pathway-related genes. Then, we used RNA ligase-mediated rapid amplification of cDNA ends (RLM-RACE) to obtain seven miRNAs cleaving eight catechin synthesis pathway-related genes including chalcone synthase (CHS), chalcone isomerase (CHI), dihydroflavonol 4-reductase (DFR), anthocyanidin reductase (ANR), leucoanthocyanidin reductase (LAR), and flavanone 3-hydroxylase (F3H). An expression analysis of miRNAs and target genes revealed that miR529d and miR156g-3p were negatively correlated with their targets CHI and F3H, respectively. The expression of other miRNAs was not significantly related to their target genes in the catechin synthesis pathway. The RLM-RACE results suggest that catechin synthesis is regulated by miRNAs that can cleave genes involved in catechin synthesis.