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Cloning of transcription factor PcFBA-1 in Pogostemon cabin and its interaction with FPPS promoter / 中国中药杂志
China Journal of Chinese Materia Medica ; (24): 412-418, 2022.
Article in Chinese | WPRIM | ID: wpr-927983
ABSTRACT
Farnesyl diphosphate synthase(FPPS) is a key enzyme at the branch point of the sesquiterpene biosynthetic pathway, but there are no reports on the transcriptional regulation of FPPS promoter in Pogostemon cabin. In the early stage of this study, we obtained the binding protein PcFBA-1 of FPPS gene promoter in P. cabin. In order to explore the possible mechanism of PcFBA-1 involved in the regulation of patchouli alcohol biosynthesis, this study performed PCR-based cloning and sequencing analysis of PcFBA-1, analyzed the expression patterns of PcFBA-1 in different tissues by fluorescence quantitative PCR and its subcellular localization using the protoplast transformation system, detected the binding of PcFBA-1 protein to the FPPS promoter in vitro with the yeast one-hybrid system, and verified its transcriptional regulatory function by dual-luciferase reporter gene assay. The findings demonstrated that the cloned PcFBA-1 had an open reading frame(ORF) of 1 131 bp, encoding a protein of 376 amino acids, containing two conserved domains named F-box-like superfamily and FBA-1 superfamily, and belonging to the F-box family. Moreover, neither signal peptide nor transmembrane domain was contained, implying that it was an unstable hydrophilic protein. In addition, as revealed by fluorescence quantitative PCR results, PcFBA-1 had the highest expression in leaves, and there was no significant difference in expression in roots or stems. PcFBA-1 protein was proved mainly located in the cytoplasm. Furthermore, yeast one-hybrid screening and dual-luciferase reporter gene assay showed that PcFBA-1 was able to bind to FPPS promoter both in vitro and in vivo to enhance the activity of FPPS promoter. In summary, this study identifies a new transcription factor PcFBA-1 in P. cabin, which directly binds to the FPPS gene promoter to enhance the promoter activity. This had laid a foundation for the biosynthesis of patchouli alcohol and other active ingre-dients and provided a basis for metabolic engineering and genetic improvement of P. cabin.
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Full text: Available Index: WPRIM (Western Pacific) Main subject: Transcription Factors / Amino Acid Sequence / Cloning, Molecular / Geranyltranstransferase / Pogostemon Type of study: Prognostic study Language: Chinese Journal: China Journal of Chinese Materia Medica Year: 2022 Type: Article

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Full text: Available Index: WPRIM (Western Pacific) Main subject: Transcription Factors / Amino Acid Sequence / Cloning, Molecular / Geranyltranstransferase / Pogostemon Type of study: Prognostic study Language: Chinese Journal: China Journal of Chinese Materia Medica Year: 2022 Type: Article