ISHS Acta Horticulturae 526: V International Symposium on Grapevine Physiology
ENZYMES OF AMMONIA ASSIMILATION IN VITIS VINIFERA L. |
| Authors: | K. A. Loulakakis, K. A. Roubelakis-Angelakis |
| Keywords: | cDNA cloning, gene expression, glutamate dehydrogenase, glutamate synthase, glutamine synthetase, grapevine, nitrogen metabolism |
| Abstract:
In an effort to better understand the physiological role of the enzymes glutamine synthetase (GS), glutamate synthase (GOGAT) and glutamate dehydrogenase (GDH) in ammonia assimilation in higher plants, we have studied their protein and molecular characteristics in an economically important perennial plant species, the grapevine. Grapevine GDH has a hexameric structure consisting of two subunit polypeptides which are associated in an ordered ratio. All isoenzymes have similar anabolic and catabolic activities and substrate kinetics. Presence of ammonium ions as nitrogen source in the culture medium of grapevine calluses resulted in increased NADH-GDH specific activity due to de novo synthesis of the  -subunit of GDH and the assembly of the more anodic isoenzymes.
Grapevine GS was analyzed into two distinct classes of isoenzymes, the cytosolic and the chloroplastic one.
A full cDNA clone encoding GDH, three structurally distinct full clones encoding cytosolic GS and two partial cDNAs encoding Fd-GOGAT were isolated from a grapevine cell suspension cDNA library and characterized.
The gene organization and the expression characteristics of the genes corresponding to the six cDNAs were determined by DNA and RNA blots, respectively.
The transcripts corresponding to these cDNAs showed differential accumulation in the various grapevine tissues tested.
Culture of cell suspensions and plantlets in media containing modified nitrogen source resulted in differential expression of the GDH, GS and Fd-GOGAT genes.
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