Identification and Characterization of Transcription Factors Regulating Terpenoid Indole Alkaloid Biosynthesis in Catharanthus roseus

Sanjay K. Singh, Barunava Patra, Joshua J. Singleton, Yongliang Liu, Priyanka Paul, Xueyi Sui, Nitima Suttipanta, Sitakanta Pattanaik, Ling Yuan

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

Abstract

Biosynthesis of the therapeutically valuable terpenoid indole alkaloids (TIAs), in the medicinal plant Catharanthus roseus, is one of the most elaborate and complex metabolic processes. Although genomic and transcriptomic resources have significantly accelerated gene discovery in the TIA pathway, relatively few genes of transcription factors (TFs) have been identified and characterized thus far. Systematic identification of TFs and elucidation of their functions are crucial for understanding TIA pathway regulation. The successful discovery of TFs in the TIA pathway has relied mostly on three different approaches, (1) identification of cis-regulatory motifs (CRMs) present in the pathway gene promoters as they often provide clues on potential TFs that bind to the promoters, (2) co-expression analysis, based on the assumption that TFs regulating a metabolic or developmental pathway exhibit similar spatiotemporal expression as the pathway genes, and (3) isolation of homologs of TFs known to regulate structurally similar or diverse specialized metabolites in different plant species. TFs regulating TIA pathway have been isolated using either an individual or a combination of the three approaches. Here we describe transcriptome-based coexpression analysis and cis-element determination to identify TFs in C. roseus. In addition, we describe the protocols for generation of transgenic hairy roots, Agrobacterium infiltration of flowers, and electrophoretic mobility shift assay (EMSA). The methods described here are useful for the identification and characterization of potential TFs involved in the regulation of special metabolism in other medicinal plants.

Original languageEnglish
Title of host publicationMethods in Molecular Biology
Pages203-221
Number of pages19
DOIs
StatePublished - 2022

Publication series

NameMethods in Molecular Biology
Volume2505
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

Bibliographical note

Publisher Copyright:
© 2022, The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.

Keywords

  • Agrobacterium rhizogenes
  • Catharanthus roseus
  • Co-expression
  • Electrophoretic mobility shift assay (EMSA)
  • Hairy roots
  • RNA-seq
  • Terpenoid indole alkaloids (TIAs)
  • Transcription factor (TF)
  • cis-regulatory element

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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