Initial organic products of assimilation of ammonium and nitrate by tobacco cells cultured on different sources of nitrogen

Glutamine is the first major organic product of assimilation of ^{13}\text{NH}{}_{4}^{+}$ by tobacco (Nicotiana tabacum L. cv. Xanthi) cells cultured on nitrate, urea, or ammonium succinate as the sole source of nitrogen, and of ^{13}\text{NO}{}_{3}^{-}$ by tobacco cells cultured on nitrate. The percentage of organic ^{13}\text{N}$ in glutamate, and subsequently, alanine, increases with increasing periods of assimilation. ^{13}\text{NO}{}_{3}^{-}$, used for the first time in a study of assimilation of nitrogen, was purified by new preparative techniques. During pulse-chase experiments, there is a decrease in the percentage of ^{13}\text{N}$ in glutamine, and a concomitant increase in the percentage of ^{13}\text{N}$ in glutamate and alanine. Methionine sulfoximine inhibits the incorporation of ^{13}\text{N}$ from ^{13}\text{NH}{}_{4}^{+}$ into glutamine more extensively than it inhibits the incorporation of ^{13}\text{N}$ into glutamate, with cells grown on any of the three sources of nitrogen. Azaserine inhibits glutamate synthesis extensively when ^{13}\text{NH}{}_{4}^{+}$ is fed to cells cultured on nitrate. These results indicate that the major route for assimilation of ^{13}\text{NH}{}_{4}^{+}$ is the glutamine synthetase-glutamate synthase pathway, and that glutamate dehydrogenase also plays a role, but a minor one. Methionine sulfoximine inhibits the incorporation of ^{13}\text{N}$ from ^{13}\text{NO}{}_{3}^{-}$ into glutamate more strongly than it inhibits the incorporation of ^{13}\text{N}$ into glutamine, suggesting that the assimilation of ^{13}\text{NH}{}_{4}^{+}$ derived from ^{13}\text{NO}{}_{3}^{-}$ may be mediated solely by the glutamine synthetase-glutamate synthase pathway.