|Gantt, Stephen - UNIVERSITY OF MINNESOTA|
Submitted to: United States of America and Mexico Symposium
Publication Type: Abstract Only
Publication Acceptance Date: November 9, 1995
Publication Date: N/A
Technical Abstract: The nodule enzymes phosphoenolpyruvate carboxylase (PEPC, EC 220.127.116.11) and NADH glutamate synthase (NADH GOGAT, EC 18.104.22.168) play pivotal roles in nitrogen (N) and carbon (C) metabolism. We have isolated and characterized the alfalfa genes encoding the nodule enhanced forms of PEPC and NADH GOGAT. PEPC comprises about 2 to 3% of total root nodule soluble protein, while NADH GOGAT is much less prevalent, comprising less than 0.25%. During effective root nodule development, PEPC and NADH GOGAT transcripts increase some 10 to 20 fold. Increased transcript accumulation is associated with two events, the first is nodule organogenesis, while the second is induction of nitrogenase activity. Ineffective nodules show expression of PEPC, but transcript levels are reduced by 60%. By comparison, NADH GOGAT transcripts are detectable in ineffective nodules at only root background levels. In situ hybridization, analysis of promoter::GUS reporter gene expression in transgenic alfalfa, and immunocytochemistry were used to assess the cellular localization and role of PEPC and NADH GOGAT in alfalfa root nodule metabolism. Promoter::GUS reporter gene constructs for both PEPC and NADH GOGAT target B glucuronidase enzyme activity to infected and uninfected cells. However, activity appears to be higher in the infected cells. Of all the enzymes we have evaluated to date, only NADH GOGAT expression appears to have an absolute requirement for some factor(s) associated with effective nodules. We postulate that NADH GOGAT is the limiting step in nodule N metabolism. Whole plant selection studies show that PEPC and NADH GOGAT are highly correlated with nitrogenase activity and can be modified by traditional plant breeding strategies.