Abstract
BACKGROUND: The relationship between gene family expansion and the resulting changes in plant phenotypes has shown remarkable complexity during the evolution. The gene family expansion has contributed to the diversity in plant phenotypes, specifically metabolites through neo-functionalization and sub-functionalization. However, the negative regulatory effects associated with the gene family expansion remain poorly understood. RESULTS: Here, we present the chromosome-scale genomes of Tinospora crispa and Tinospora sinensis. Comparative genomic analyses demonstrated conserved chromosomal evolution within the Menispermaceae family. KEGG analysis revealed a significant enrichment of genes related to terpenoid biosynthesis in T. sinensis. However, T. crispa exhibited a higher abundance of terpenoids compared to T. sinensis. Detailed analysis revealed the expansion of genes encoding 1-hydroxy-2-methyl 2-(E)-butenyl 4-diphosphate synthase (HDS), a key enzyme in the 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway of terpenoid biosynthesis in T. sinensis. TsiHDS4 retained the ancestral function of converting methylerythritol cyclic diphosphate (MEcPP) to (E)-4-hydroxy-3-methylbut-2-enyl diphosphate (HMBPP). However, the noncanonical CDS-derived small peptide TsiHDS5 was shown to interact with TsiHDS4, inhibiting its catalytic activity. This interaction reduced the levels of HMBPP and isopentenyl pyrophosphate (IPP), which represent key substrates for downstream terpenoid biosynthesis. CONCLUSIONS: These findings offer clues to decipher the variations in the MEP pathway of terpenoid biosynthesis between T. crispa and T. sinensis and form a basis for further detailed research on the negative regulation of expanded genes.