Abstract
In Catharanthus roseus cells, auxins are known to negatively regulate the biosynthesis of monoterpenoid indole alkaloids (MIA), a class of valuable secondary metabolites. Despite extensive studies of this regulation, no protein of the auxin signaling pathway has been isolated to date in this plant. We therefore decided to clone and characterize a C. roseus Aux/IAA protein that belongs to a family of gene expression repressors mediating auxin effects. Using PCR, a cDNA encoding the first C. roseus Aux/IAA was cloned and named CrIAA1. The deduced amino acid sequence has four highly conserved domains that are typical of the Aux/IAA protein family and has high homology to the Aux/IAA isoforms of Arabidopsis (>67%). The CrIAA1 gene expression, monitored by real-time PCR, was found to be dramatically induced by auxin treatment in C. roseus cells. Using GFP imagery and a bimolecular fluorescence complementation assay, we found that CrIAA1 can form oligomers in the nucleus. We also found that CrIAA1 is quickly degraded following auxin treatments, suggesting that auxin regulates CrIAA1 availability via a feedback mechanism. These results should help to elucidate the molecular nature of the processes responsible for the auxin-mediated regulation of MIA biosynthesis in C. roseus.
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Acknowledgments
This research was financially supported by the Ministère de l’Enseignement Supérieur et de la Recherche. We thank Dr. A. J. Simkin, who kindly provided the C. roseus cDNA library, and Vic Norris for help with the English. G. Guirimand was financed by a MENRT fellowship.
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344_2010_9187_MOESM1_ESM.tif
Supplementary Fig. 1 Control of the nuclear localization of the BiFC complexes displayed in Fig. 3B. C. roseus cells were transiently cotransformed with BiFC vectors (CrIAA1 or Arabidopsis bZIP63 transcription factor as a positive control) as labeled on the top (fusion with YFPN fragment) and on the left (fusion with the YFPC fragment) in combination with the CFP-GUS-nucleoplasmin NLS-CFP marker. The CFP channel and DIC images are in the same focal plane as for the cells analyzed by BiFC in Fig. 3B. These results demonstrate that BiFC complexes colocalize with nuclear CFP marker and that the cells are transformed even in the absence of BiFC complex formation. Scale bar = 10 μm (TIF 6799 kb)
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Poutrain, P., Guirimand, G., Glévarec, G. et al. Molecular Characterization of an Aux/IAA of Catharanthus roseus . J Plant Growth Regul 30, 235–241 (2011). https://doi.org/10.1007/s00344-010-9187-3
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DOI: https://doi.org/10.1007/s00344-010-9187-3