title: The basic helix-loop-helix transcription factor BIS2 is essential for monoterpenoid indole alkaloid production in the medicinal plant Catharanthus roseus.
abstract: Monoterpenoid indole alkaloids (MIAs) are produced as plant defence compounds. In the medicinal plant Catharanthus roseus, they comprise the anticancer compounds vinblastine and vincristine. The iridoid (monoterpenoid) pathway forms one of the two branches that feed MIA biosynthesis and its activation is regulated by the transcription factor (TF) basic helix-loop-helix (bHLH) iridoid synthesis 1 (BIS1). Here, we describe the identification and characterisation of BIS2, a jasmonate (JA)-responsive bHLH TF expressed preferentially in internal phloem-associated parenchyma cells, which transactivates promoters of iridoid biosynthesis genes and can homodimerise or form heterodimers with BIS1. Stable overexpression of BIS2 in C. roseus suspension cells and transient ectopic expression of BIS2 in C. roseus petal limbs resulted in increased transcript accumulation of methylerythritol-4-phosphate and iridoid pathway genes, but not of other MIA genes or triterpenoid genes. Transcript profiling also indicated that BIS2 expression is part of an amplification loop, as it is induced by overexpression of either BIS1 or BIS2. Accordingly, silencing of BIS2 in C. roseus suspension cells completely abolished the JA-induced upregulation of the iridoid pathway genes and subsequent MIA accumulation, despite the presence of induced BIS1, indicating that BIS2 is essential for MIA production in C. roseus.
authors.affiliation: Department of Plant Systems Biology, VIB, Technologiepark 927, B-9052, Gent, Belgium.|Department of Plant Biotechnology and Bioinformatics, Ghent University, Technologiepark 927, B-9052, Gent, Belgium.
authors.forename: Alex
authors.initials: A
authors.lastname: Van Moerkercke
authors.affiliation: Institute of Biology, Leiden University, P.O. Box 9505, 2300 RA, Leiden, The Netherlands.
authors.forename: Priscille
authors.initials: P
authors.lastname: Steensma
authors.affiliation: Institute of Biology, Leiden University, P.O. Box 9505, 2300 RA, Leiden, The Netherlands.
authors.forename: Ivo
authors.initials: I
authors.lastname: Gariboldi
authors.affiliation: Institute of Biology, Leiden University, P.O. Box 9505, 2300 RA, Leiden, The Netherlands.
authors.forename: Javiera
authors.initials: J
authors.lastname: Espoz
authors.affiliation: Institute of Biology, Leiden University, P.O. Box 9505, 2300 RA, Leiden, The Netherlands.
authors.forename: Purin C
authors.initials: PC
authors.lastname: Purnama
authors.affiliation: Department of Plant Systems Biology, VIB, Technologiepark 927, B-9052, Gent, Belgium.|Department of Plant Biotechnology and Bioinformatics, Ghent University, Technologiepark 927, B-9052, Gent, Belgium.
authors.forename: Fabian
authors.initials: F
authors.lastname: Schweizer
authors.affiliation: Department of Plant Systems Biology, VIB, Technologiepark 927, B-9052, Gent, Belgium.|Department of Plant Biotechnology and Bioinformatics, Ghent University, Technologiepark 927, B-9052, Gent, Belgium.
authors.forename: Karel
authors.initials: K
authors.lastname: Miettinen
authors.affiliation: Department of Plant Systems Biology, VIB, Technologiepark 927, B-9052, Gent, Belgium.|Department of Plant Biotechnology and Bioinformatics, Ghent University, Technologiepark 927, B-9052, Gent, Belgium.
authors.forename: Robin
authors.initials: R
authors.lastname: Vanden Bossche
authors.affiliation: Department of Plant Systems Biology, VIB, Technologiepark 927, B-9052, Gent, Belgium.|Department of Plant Biotechnology and Bioinformatics, Ghent University, Technologiepark 927, B-9052, Gent, Belgium.
authors.forename: Rebecca
authors.initials: R
authors.lastname: De Clercq
authors.affiliation: Institute of Biology, Leiden University, P.O. Box 9505, 2300 RA, Leiden, The Netherlands.
authors.forename: Johan
authors.initials: J
authors.lastname: Memelink
authors.affiliation: Department of Plant Systems Biology, VIB, Technologiepark 927, B-9052, Gent, Belgium. algoo@psb.vib-ugent.be.|Department of Plant Biotechnology and Bioinformatics, Ghent University, Technologiepark 927, B-9052, Gent, Belgium. algoo@psb.vib-ugent.be.
authors.forename: Alain
authors.initials: A
authors.lastname: Goossens
mesh_terms: D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics / Q000378:metabolism*; D030008:Catharanthus / Q000235:genetics / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant / Q000235:genetics / Q000502:physiology; D026121:Indole Alkaloids / Q000378:metabolism*; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D010946:Plants, Medicinal / Q000235:genetics / Q000378:metabolism*
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D026121:Indole Alkaloids; D010940:Plant Proteins
keywords: 
  Catharanthus roseus
  ; Madagascar periwinkle; alkaloid; basic helix-loop-helix; iridoid; jasmonate; monoterpenoid; terpenoid