Enzyme Inhibitor Studies Reveal Complex Control of Methyl-D-Erythritol 4-Phosphate (MEP) Pathway Enzyme Expression in Catharanthus roseus
Figure 7
Effect of fosmidomycin on the expression of DXS protein and DXS isoform (1, 3) transcripts in 8-day-old seedlings of Arabidopsis thaliana.
A. thaliana Ler seedlings were cultivated for 8 days at 22°C in petri dishes on filter paper as described by Guevara-Garcia et al. [23]. Light period was 16 hrs (230 µmol m−2 s−1). Thereafter, 5 ml of a 100 µM fosmidomycin solution (or water as control) were added. After the indicated time intervals, seedlings were harvested and processed for MEP pathway protein and transcript analysis, respectively. (A) DXS, DXR, and HDS proteins were detected by immunoblot with polyclonal antisera raised against recombinant C. roseus DXS2A, DXR and HDS proteins. Note that exposure time was five times longer for Arabidopsis samples than for C. roseus samples. (B) transcript amounts for DXS isoforms (1& 3) were determined by qPCR. Relative transcript levels were calculated by normalizing to actin as reference gene. Means of triplicate values from a representative experiment are presented ± SD.