Understanding in vivo carbon precursor supply for fatty acid synthesis in leaf tissue

The principal supply of carbon precursors for fatty acid synthesis in leaf tissue has been a much debated topic, with some experiments suggesting a di rect supply from the C3 products of photosynthetic carbon fixation and othe rs suggesting the utilization of free acetate (for which concentrations in leaves in the range of 0.05-1.4 mM have been reported). To address this iss ue we first reassessed the in vivo rate of fatty acid synthesis using a new method, that of [C-13]carbon dioxide labeling of intact Arabidopsis plants with the subsequent analysis of fatty acids by gas chromatography-mass spe ctrometry (GC-MS). This method gave an average value of 2.3 mmoles carbon a toms h(-1) mg chlorophyll(-1) for photosynthetic tissues. The method was ex tended by isotopic dilution analysis to measure the rate of fatty acid synt hesis in the dark. There was negligible fatty acid synthesis (<5% of the ra te in the light) in the dark. In addition, the method allowed an estimate o f the absolute rate of fatty acid degradation of about 4% of the total fatt y acid content per day. With the in vivo rate of fatty acid synthesis in th e light defined, if the bulk tissue acetate concentration available for fat ty acid synthesis is 1 mM, this acetate pool can sustain fatty acid synthes is for approximately 60 min. When the leaves of Arabidopsis, barley and pea were given a 5 min pulse of [C-14]carbon dioxide, the label rapidly appear ed in fatty acids with a lag phase of less than 2-3 min. Continuous labelin g with [C-14]carbon dioxide, for up to 1 h, showed a similar result. Furthe rmore, C-14-label in free acetate was less than 5% of that in fatty acids. In conclusion, these data suggest that either the bulk pool of acetate is n ot involved in fatty acid synthesis or the concentration of acetate must be less than 0.05 mM under strong illumination.