Unravelling lncRNA mediated gene expression as potential mechanism for regulating secondary metabolism in Citrus limon

Abstract Plant long non-coding RNAs (lncRNAs) are emerging as important regulators of gene expression through various mechanisms viz. binding to double-stranded DNA (dsDNA), physically targeting mRNAs, acting as microRNA sponges. Characteristic secondary metabolites and their content determine the market value of Citrus plant products for culinary and industrial uses. Though lncRNAs are emerging as important molecular players that regulate secondary metabolites in plants but the roles of lncRNAs are yet unknown for the regulation of secondary metabolites in Citrus limon. In this study, a total of 11814 lncRNAs were identified from four RNA-seq datasets comprising of data from four different tissues of C. limon viz. leaf, bud, fruit and peel. Co-expression analysis of whole set of genes with identified lncRNAs revealed a total of 632 lncRNAs correlating with 5810 genes. Functional annotation of genes correlating with lncRNAs unveiled the association of 379 genes belonging to the biosynthesis of secondary metabolite pathways and among them, 36 and 12 genes are from terpenoids and flavonoids biosynthesis pathways respectively. Expression correlation analysis unfolded the possible role of 113 lncRNAs in terpenoids metabolism and 29 lncRNAs in the flavonoids metabolic pathway. Seventy nine lncRNAs were predicted to take part in miRNA-mediated gene regulation, out of which 10 lncRNAs were predicted to act as endogenous target mimics (eTMs) for miRNAs and 74 lncRNAs were found to act as miRNA targets. Expression of six predicted lncRNAs and seven correlated genes involved in terpenoids and flavonoids metabolism were validated using semi-quantitative RT- PCR.