Blackwell Publishing Journal Backfiles 1879-2005
Soybean (Glycine max [L.] Merr. cv. Century) seedlings were germinated in vermiculite, sub-irrigated with a complete nutrient solution, without nitrogen or supplemented with 10 mM KNO3, 5 mM (NH4)2SO4, or 5 mM NH4NO3. After 14 days in the light, or 5 days in the dark, tissues from different organs were harvested separately. Similarly, tissues from different organs from 14- or 21-day-old nodulated or non-nodulated soybean seedlings, maintained in the absence of nitrogen, were harvested. Proteins and total RNA were isolated from the different plant organs and used for immunoblot and RNA blot analyses, respectively. Protein or RNA blots were separately incubated with antisera or hybridized with probes specific for either ferredoxin-dependent glutamate synthase (Fd-GOGAT) or NADH-dependent glutamate synthase (NADH-GOGAT). The specific activity and the abundance of the Fd-GOGAT peptide and transcript increased, approximately 3–10 times, in cotyledons and hypocotyls/stems in plants germinated in the light compared with those germinated in the dark. Fd-GOGAT activity, peptide, and transcript were highest in leaves. Except for increases in the specific activity of samples from roots treated with (NH4)2SO4 or NH4NO3, there were minor or no changes in Fd-GOGAT activity, peptide and transcript among organs of seedlings treated with different nitrogen sources or by nodulation. Low levels of NADH-GOGAT transcript were detected in all organs. NADH-GOGAT activity, peptide, and transcript increased in roots of seedlings treated with different nitrogen sources, but these changes were more apparent on RNA blots versus immunoblots. The highest NADH-GOGAT activity and most abundant amounts of the peptide and transcript were observed in nodules. Despite being induced by different environmental factors, both GOGAT activities are controlled, at least in part, by either gene expression or by RNA stability, because in most instances, both isoenzymes exhibited paralleled changes in specific activity and the abundance of their corresponding peptides and transcripts on immunoblot and RNA blots, respectively. However, there were some exceptions to the parallel increases in specific activities, peptides and transcripts which suggest that post-translational modification may also regulate the activties of the two GOGAT isoforms. Collectively, the results presented here suggest that the two GOGAT isoforms have distinct physiological functions in soybean.
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