Decreased NADH glutamate synthase activity in nodules and flowers of alfalfa (Medicago sativa L.) transformed with an antisense glutamate synthase transgene
Ma. Schoenbeck et al., Decreased NADH glutamate synthase activity in nodules and flowers of alfalfa (Medicago sativa L.) transformed with an antisense glutamate synthase transgene, J EXP BOT, 51(342), 2000, pp. 29-39
Legumes obtain a substantial portion of their nitrogen (N) from symbiotic N
-2 fixation in root nodules, The glutamine synthetase (GS, EC 6.3.1.2)/glut
amate synthase (GOGAT) cycle is responsible for the initial N assimilation.
This report describes the analysis of a transgenic alfalfa (Medicago sativ
a L.) line containing an antisense NADH-GOGAT (EC 1.4.1.14) under the contr
ol of the nodule-enhanced aspartate aminotransferase (AAT-2) promoter. In o
ne transgenic line, NADH-GOGAT enzyme activity was reduced to approximately
50%, with a corresponding reduction in protein and mRNA, The transcript ab
undance for cytosolic GS, ferredoxin-dependent GOGAT (EC 1,4,7,1), AAT-2 (E
C 2.6.1.1), asparagine synthase (EC 6.3.5,4), and phosphoenolpyruvate carbo
xylase (PEPC, EC 4,1,1.31) were unaffected, as were enzyme activities for A
AT, PEPC and GS. Antisense NADH-GOGAT plants grown under symbiotic conditio
ns were moderately chlorotic and reduced in growth and N content, even thou
gh symbiotic N-2 fixation was not significantly reduced. The addition of ni
trate relieved the chlorosis and restored growth and N content. Surprisingl
y, the antisense NADH-GOGAT plants were male sterile resulting from inviabl
e pollen, A reduction in NADH-GOGAT enzyme activity and transcript abundanc
e in the antisense plants was measured during the early stages of flower de
velopment. Inheritance of the transgene was stable and resulted in progeny
with a range of NADH-GOGAT activity, These data indicate that NADH-GOGAT pl
ays a critical role in the assimilation of symbiotically fixed N and during
pollen development.