关键词: 葡萄, TPS基因家族, 表达分析, 香气, 代谢组学

Abstract:

【Objective】Based on grape genome information, this study identifies and analyzes the terpene synthase (TPS) gene family expression in grapes using bioinformatics methods. This research lays an important foundation for subsequent studies on the biological functions of VvTPSs and grape breeding. 【Method】The VvTPS gene family was identified using the protein sequences of Arabidopsis AtTPS gene family members and the reported Hidden Markov model (HMM) files PF01397 and PF03936. Various bioinformatics tools, including Expasy, Tbtools, MEME, MEGA, MCScanX, SPOMA, WoLF PSORT, and PlantCARE were used to analyze the physicochemical characteristics, phylogenetic tree, chromosomal distributions, gene structure, subcellular localization, secondary structure of protein, and cis-acting elements in the promoters of gape TPS family genes. Additionally, the expression profiles of TPS genes in two aromatic grape varieties Red Globe (neutral) and Muscat Hamburg (muscat) were analyzed using qPCR. The transgenic tomato overexpressed VvTPS4 was used to study its function in influencing the muscat aroma by metabolomics. 【Result】A total of 65 VvTPS gene family members were identified, encoding proteins ranging from 339 to 840 amino acids, with an average molecular mass of 64.13 kDa and theoretical isoelectric points of 4.93 to 7.65. Most members have 7 exon structures. Phylogenetic analysis classified the grape TPS genes into five subfamilies: TPS-a, TPS-b, TPS-c, TPS-e/f, and TPS-g. Subcellular localization analysis indicated that most VvTPS proteins are localized in chloroplasts and cytoplasm, with 10 members from the TPS-g subfamily found in plastids. Promoter analysis revealed numerous cis-acting elements associated with responses to light, temperature, drought, hormones, and defense mechanisms. Of the 29 VvTPS genes cloned, most showed higher expression levels in the muscat-flavored grape variety compared to the non-aromatic variety. In transgenic tomatoes overexpressing VvTPS4, volatile monoterpenoids were significantly accumulated, with linalool levels increasing 20.73-fold and L-α-terpineol levels rising 14.55-fold compared to the wild type. Flavor characteristics analysis demonstrated these two compounds have floral aroma and are the main characteristic substances affecting the aroma of muscat fragrance.【Conclusion】Sixty-five VvTPS genes were identified in grapes, showing high conservation with some variations. Several TPS genes showed significantly expressed in muscat-flavored grape varieties. Overexpression of VvTPS4 led to a significant increase the accumulation of volatile monoterpenoids, which may play an essential role in developing the muscat aroma in grapes.

Key words: grape, TPS gene family, expression analysis, aroma, metabolomics