Whole-genome,transcriptome,and methylome analyses provide insights into the evolution of platycoside biosynthesis in Platycodon grandiflorus,a medicinal plant

Triterpenoid saponins(TSs)are common plant defense phytochemicals with potential pharmaceutical properties.Platycodon grandiflorus(Campanulaceae)has been traditionally used to treat bronchitis and asthma in East Asia.The oleanane-type TSs,platycosides,are a major component of the P.grandiflorus root extract.Recent studies show that platycosides exhibit anti-inflammatory,antiobesity,anticancer,antiviral,and antiallergy properties.However,the evolutionary history of platycoside biosynthesis genes remains unknown.In this study,we sequenced the genome of P.grandiflorus and investigated the genes involved in platycoside biosynthesis.The draft genome of P.grandiflorus is 680.1 Mb long and contains 40,017 protein-coding genes.Genomic analysis revealed that the CYP716 family genes play a major role in platycoside oxidation.The CYP716 gene family of P.grandiflorus was much larger than that of other Asterid species.Orthologous gene annotation also revealed the expansion ofβ-amyrin synthases(bASs)in P.grandiflorus,which was confirmed by tissue-specific gene expression.In these expanded gene families,we identified key genes showing preferential expression in roots and association with platycoside biosynthesis.In addition,wholegenome bisulfite sequencing showed that CYP716 and bAS genes are hypomethylated in P.grandiflorus,suggesting that epigenetic modification of these two gene families affects platycoside biosynthesis.Thus whole-genome,transcriptome,and methylome data of P.grandiflorus provide novel insights into the regulation of platycoside biosynthesis by CYP716 and bAS gene families.

Molecular Characterization of the Calvin Cycle Enzyme Phosphoribulokinase in the Stramenopile Alga Vaucheria litorea and the Plastid Hosting Mollusc Elysia chlorotica

Phosphoribulokinase （PRK）, a nuclear-encoded plastid-localized enzyme unique to the photosynthetic carbon reduction （Calvin） cycle, was cloned and characterized from the stramenopile alga Vaucheria litorea. This alga is the source of plastids for the mollusc （sea slug） Elysia chlorotica which enable the animal to survive for months solely by photoautotrophic CO2 fixation. The 1633-bp V. litorea prk gene was cloned and the coding region, found to be interrupted by four introns, encodes a 405-amino acid protein. This protein contains the typical bipartite target sequence expected of nuclearencoded proteins that are directed to complex （i.e. four membrane-bound） algal plastids. De novo synthesis of PRK and enzyme activity were detected in E. chlorotica in spite of having been starved of V. litorea for several months. Unlike the algal enzyme, PRK in the sea slug did not exhibit redox regulation. Two copies of partial PRK-encoding genes were isolated from both sea slug and aposymbiotic sea slug egg DNA using PCR. Each copy contains the nucleotide region spanning exon 1 and part of exon 2 of V litorea prk, including the bipartite targeting peptide. However, the larger prk fragment also includes intron 1. The exon and intron sequences of prk in E. chlorotica and V/itorea are nearly identical. These data suggest that PRK is differentially regulated in V. litorea and E. chlorotica and at least a portion of the V. litorea nuclear PRK gene is present in sea slugs that have been starved for several months.