Nitrogen retrieval and assimilation by ectomycorrhizal fungi is thought to play a central role in the economy of the symbiotic interaction. Here, we report on the molecular characterization of the key N assimilation enzyme glutamine synthetase from the mycorrhizal ascomycete Tuber borchii (TbGS). TbGS displayed a strong positive co-operativity (n=1.7+/-0.29) and an unusually high S0,5 value (54+/-16mM; S0,5 is the substrate concentration value at which v=1/2 Vmax) for glutamate, and a correspondingly low sensitivity towards inhibition by the glutamate analogue herbicide phosphinothricin. T h e T b G S m R N A, which is encoded by a single-copy gene in the Tuber genome, was upregulated in N starved mycelia and returned to basal levels upon resupplementation of various forms of N, the most effective of which was nitrate. Both responses were accompanied by parallel variations of TbGS protein amount and glutamine synthetase activity, indicating that TbGS levels are primarily controlled at the pretranslational level. As revealed by a comparative analysis of TbGS and of the mRNAs for the metabolically related enzymes glutamate dehydrogenase and glutamate synthase, TbGS is not only the sole messenger that positively responds to N starvation, but also the most abundant under N limiting conditions. A similar, but even more discriminating expression pattern, with practically undetectable GDH mRNA levels, was observed in fruitbodies. The TbGS mRNA was also found to be expressed in symbiosis-engaged hyphae, with higher hybridation signals in hyphae that were penetrating among and within root cells.
Distinctive functional properties and expression profiles of glutamine synthetase from a plant symbiotic fungus
Balestrini R;
2003
Abstract
Nitrogen retrieval and assimilation by ectomycorrhizal fungi is thought to play a central role in the economy of the symbiotic interaction. Here, we report on the molecular characterization of the key N assimilation enzyme glutamine synthetase from the mycorrhizal ascomycete Tuber borchii (TbGS). TbGS displayed a strong positive co-operativity (n=1.7+/-0.29) and an unusually high S0,5 value (54+/-16mM; S0,5 is the substrate concentration value at which v=1/2 Vmax) for glutamate, and a correspondingly low sensitivity towards inhibition by the glutamate analogue herbicide phosphinothricin. T h e T b G S m R N A, which is encoded by a single-copy gene in the Tuber genome, was upregulated in N starved mycelia and returned to basal levels upon resupplementation of various forms of N, the most effective of which was nitrate. Both responses were accompanied by parallel variations of TbGS protein amount and glutamine synthetase activity, indicating that TbGS levels are primarily controlled at the pretranslational level. As revealed by a comparative analysis of TbGS and of the mRNAs for the metabolically related enzymes glutamate dehydrogenase and glutamate synthase, TbGS is not only the sole messenger that positively responds to N starvation, but also the most abundant under N limiting conditions. A similar, but even more discriminating expression pattern, with practically undetectable GDH mRNA levels, was observed in fruitbodies. The TbGS mRNA was also found to be expressed in symbiosis-engaged hyphae, with higher hybridation signals in hyphae that were penetrating among and within root cells.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
