Researchers have successfully generated a high-quality, haplotype-resolved pentaploid genome of Gynostemma pentaphyllum, a plant species that serves as an important natural source of dammarane-type triterpenoid saponins with applications in healthcare and pharmaceutical industries. The study, published in Engineering, provides new insights into chromosomal evolution and reveals a case of convergent evolution in the biosynthesis of protopanaxadiol (PPD)-type saponins.

The research team focused on the Jiangxi variety of G. pentaphyllum, which is rich in PPD-type saponins, in contrast to the previously reported Anhui variety. Karyotype analysis revealed the sequenced material to be pentaploid, with a k-mer-based evaluation indicating a total genome size of 2.1 Gb. The final assembly, designated GpJX, comprised 55 chromosomes organized into 11 homologous groups, with five chromosomes per group, and was 2.48 Gb in size. The assembly achieved a BUSCO score of 99.0% for base-level completeness, an LTR Assembly Index of 11.3, and a quality value of 60.7 with 98.7% k-mer completeness.

Through comparative genomic analyses, the researchers reconstructed and updated the ancestral karyotype of Cucurbitaceae to include 14 chromosomes, using Momordica charantia as the reference species. Analysis revealed that compared with Benincasa hispida (K_s = 1.27), M. charantia (K_s = 1.22) has a slower evolutionary rate, suggesting better preservation of ancestral chromosomal characteristics. The study identified eight shared chromosome-like synteny blocks, exceeding the seven previously identified in B. hispida.

The research revealed extensive chromosomal inversions and notable sequence variation in centromeric regions among G. pentaphyllum accessions of different ploidy levels. Statistical analyses revealed hundreds of inversion variants between the diploid GpT2T and each GpJX haplotype, with 32 inversion variants exceeding 1 Mb identified between the two genomes. The researchers hypothesize that transposable elements played a key role in shaping chromosomal structure and centromere evolution, potentially contributing to ploidy diversification in G. pentaphyllum.

A significant finding of the study was the characterization of two PPD synthases (PPDSs) from the CYP88 family of cytochrome P450s. In G. pentaphyllum, the hydroxylation of dammarenediol-II to produce PPD represents the only unknown step in the biosynthesis of PPD-type saponins. The researchers identified GpCYP88G1 and GpCYP88G2, which catalyze the C12 hydroxylation of dammarenediol-II to produce PPD. Notably, CYP716A47 from Panax ginseng has been functionally characterized as a PPD synthase belonging to the CYP716 family, while the homologous gene in G. pentaphyllum, designated CYP716_89, exhibited the highest sequence similarity (48%) to CYP716A47 but was unable to catalyze the conversion.

Molecular docking analysis revealed that, compared with the PPDS of P. ginseng, the isozyme in G. pentaphyllum orients its substrate in the opposite direction during catalysis because of distinct amino acid interactions. Phylogenetic analysis further indicated that the PPDSs in G. pentaphyllum and P. ginseng independently recruited different key residues, highlighting a case of convergent evolution. The sequence similarity between GpCYP88G1/2 in G. pentaphyllum and CYP716A47 in P. ginseng was only 23.87%, suggesting a distant relationship between the two genes.

The researchers conclude that the high-quality genome assembly provides new insights into chromosome evolution and the mechanisms underlying ploidy diversification while also establishing a foundation for advancing understanding of triterpene saponin biosynthesis in this species.

The paper “The Haplotype-Resolved Pentaploid Gynostemma pentaphyllum Genome Provides Insights into Chromosomal Evolution and the Convergent Evolution of Protopanaxadiol Synthases,” is authored by Chuyi Zhang, Lingling Yun, Ziqin Li, Sijie Sun, Yini Niu, Li Qiu, Feng Cao, Xiaofeng Shen, Li Xiang, Ying Li, Baolin Guo, Vincent Courdavault, Chao Sun. Full text of the open access paper: https://doi.org/10.1016/j.eng.2025.11.022. For more information about Engineering, visit the website at https://www.sciencedirect.com/journal/engineering.