Comparative Transcriptome Analysis Reveals Genes Associated with Alkaloid Diversity in Javanese Long Pepper (Piper retrofractum) Fruits

Alkaloids are a class of secondary metabolites that play multifaceted roles in plant physiology, including defense mechanisms and interactions with other organisms. The alkaloids from Piper retrofractum (Javanese long pepper) fruits offer potential alternatives to synthetic pesticides due to their natural origin and insecticide properties. However, information on particular alkaloid biosynthesis pathways is required to enhance individual alkaloid production via metabolic engineering. Here, we perform HPLC profiling to demonstrate that fruit ripening influences the alkaloid diversity in P. retrofractum. De novo transcriptomic profiling of young, green mature, and red ripened fruits revealed that the piperine biosynthesis pathway genes were highly upregulated in the mature fruits. However, an enhanced accumulation of methyl piperate and guineensine in the ripened fruit was observed, entailing ripening-related differential gene expression to synchronize the alkaloid biosyntheses. Gene expression clustering and functional enrichment analysis identified a large group of genes involved in diverse biosynthetic processes explicitly enriched in the ripened fruits. A cohort of genes encoding for “Alkaloid Biosynthesis”, remarkably upregulated in the ripening fruits, indicates they may function directly in alkaloid diversity during a later stage of fruit development. This study provides the basis for metabolic engineering to enhance alkaloid diversity and production.