ASSIMILATION OF NITRATE AND AMMONIUM BY SULFUR DEFICIENT ZEA-MAYS CELLS

In plants, mineral limitations including sulphur deficiency, often res ult in the accumulation of free amino acids. These could come from pro tein breakdown or from de novo synthesis. To decide between these alte rnatives cell suspension cultures of maize (Zea mays L.) growing in a modified Murashige and Skoog medium containing 1/10 of the normal nitr ogen were subjected to sulphate starvation for 4 or 11 d. Analysis of amino acids by High Performance Liquid Chromatography (HPLC) showed a net increase with time of sulphate deprivation in the pools of glutami ne (gin), asparagine (asn), glutamate (glu), aspartate (asp) and alani ne (ala). In batches of cells grown with or without sulphate for 4 or 11 d, the incorporation of (NO3-)-N-15 and (NH4+)-N-15 during short pe riods (10-80 min) was analysed by Gas Chromatography-Mass Spectrometry (GC-MS). In -S cultures after 11 days a much smaller proportion of th e gin molecules in the large gin pool was labelled in comparison with the cells from +S cultures. But the amount of N-15-gln formed during t he labelling period was greater on day 11 than on the day 4. In vivo N -15-Nuclear Magnetic Resonance (NMR) spectroscopy was used to follow t he assimilation of N-15 from (NH4NO3)-N-15 to amino compounds over a 4 h period. During this time the energy status and the internal pH of t he cells were monitored by P-31-NMR. Control (+S) cell suspensions gav e N-15 spectra where amino acid signals showed very similar relative c oncentrations. In spectra from -S cells there was a greater signal of the amide-N of glutamine than in controls. These results indicate that amino acid accumulation is a consequence of de novo synthesis, not of protein breakdown.