Catharanthus roseus G-box binding factors 1 and 2 act as repressors of strictosidine synthase gene expression in cell cultures.
The enzyme encoded by the strictosidine synthase (Str) gene catalyses a key step in the biosynthesis of therapeutically valuable terpenoid indole alkaloids. In Catharanthus roseus the Str gene was shown to be regulated by a wide variety of signals including auxin, methyl jasmonate and fungal elicitors in cell suspension cultures and by tissue-specific control in plant organs. The Str promoter contains a functional G-box (CACGTG) cis-regulatory sequence. In order to understand better the mechanisms involved in the regulation of Str gene expression, we isolated the C. roseus cDNAs encoding G-box binding factors Crgbf1 and Crgbf2. The binding specificity of their protein products CrGBF1 and CrGBF2 was analysed by competitive electrophoresis mobility and saturation binding assays. CrGBF1 had a high binding specificity for class I G-boxes including the Str G-box. CrGBF1 showed a lower affinity for class II G-boxes and for the G-box-like element (AACGTG) found in the tryptophan decarboxylase (Tdc) gene which encodes another enzyme involved in TIA biosynthesis. CrGBF2 showed a high affinity for all types of G-boxes tested and to a lesser extent for the Tdc G-box-like element. Transient bombardment experiments demonstrated that both CrGBF1 and CrGBF2 can act in vivo as transcriptional repressors of the Str promoter via direct interaction with the G-box. These data indicate that GBFs may play functional role in the regulation of expression of the terpenoid indole alkaloid biosynthetic gene Str.
Sibéril
Y
Y
Université de Tours, EA 2106, Biomolécules et Biotechnologies Végétales, UFR des Sciences et Techniques, Laboratoire de Physiologie Végétale, France.
Benhamron
S
S
Memelink
J
J
Giglioli-Guivarc'h
N
N
Thiersault
M
M
Boisson
B
B
Doireau
P
P
Gantet
P
P
D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive; D015152:Blotting, Northern; D015139:Blotting, Southern; D019759:Carbon-Nitrogen Lyases / Q000235:genetics*; D004247:DNA / Q000235:genetics / Q000378:metabolism; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D018744:DNA, Plant / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D051790:G-Box Binding Factors; D015971:Gene Expression Regulation, Enzymologic; D018506:Gene Expression Regulation, Plant; D008969:Molecular Sequence Data; D059828:Plant Cells; D010944:Plants / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D018749:RNA, Plant / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics*; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D014018:Tissue Distribution; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism
D018076:DNA, Complementary; D018744:DNA, Plant; D004268:DNA-Binding Proteins; D051790:G-Box Binding Factors; D020033:Protein Isoforms; D018749:RNA, Plant; D012097:Repressor Proteins; D014157:Transcription Factors; D004247:DNA; D019759:Carbon-Nitrogen Lyases; C022684:strictosidine synthetase