Extended Data Fig. 6: Incorporation of [2-13C] pyruvate into intermediates of the 2-C-methylerythritol-4-phosphate (MEP) pathway in Arabidopsis seedlings (7-day old) vs. rosettes (42-day old).
From: Rubisco supplies pyruvate for the 2-C-methyl-d-erythritol-4-phosphate pathway
Following incubation with [2-13C] pyruvate, 13C was detected in MEP pathway intermediates in seedlings but not in rosette leaves. a, Label incorporation into 1-deoxy-d-xylulose-5-phosphate (DXP). b, Label incorporation into 2-C-methylerythritol-2,4,-cyclodiphosphate (MEcDP); c, Label incorporation into isopentenyl and dimethylallyl diphosphate (IDP/DMADP). IDP and DMADP, which are the products of both the MEP pathway in the chloroplast and the mevalonate pathway in the cytosol, displayed measurable label incorporation in both seedlings and rosette leaves. d, Detection of exogenously supplied [2-13C] pyruvate in extracts of seedlings or rosette leaves. Seedlings were grown in ¾ MS media supplemented (or not) with 5 mM 2-13C pyruvate for 7 days while leaves of rosette plants were infiltrated for 24 hours with MS media supplemented with labelled pyruvate at the same concentration (or MS media only for controls). Pooled leaves or seedlings were flash-frozen and extracted for LCMS/MS and GCMS analysis. Percent labelling incorporation into each metabolite was calculated using the equation (\(1/N\)) \({\sum }_{i}^{N}Mi\times i\), where N is the number of carbon atoms in the molecule and Mi is the fractional abundance of the ith isotopologue. Error bars indicate standard deviation of measurement (n = 3-4). Asterisks indicate significant differences (** P < 0.001; *** P < 0.0001) as determined by Student’s two-sided t-test. n.s., Not statistically significant.
