Significance of NADP/NAD glutamate dehydrogenase ratio in the dimorphic behavior of Benjaminiella poitrasii and its morphological mutants

Abstract

Studies on the levels of glutamate dehydrogenase (GDH), glutamine synthetase, and glutamate synthase were carried out as a function of temperature, nutritional conditions, and the morphological (yeast or mycelium) form of Benjaminiella poitrasii. Since both NAD-and NADP-dependent GDH activities were found in B. poitrasii, the quantitative relation between these two enzymes expressed as the NADP-GDH/NAD-GDH activity ratio (GDH ratio) was studied to evaluate its possible role in the morphogenesis. In the yeast-to-mycelium transition, a decrease in the GDH ratio occurred (between 1 and 2 h) and germ tube formation could be observed only at 3 h. Under similar sets of experimental conditions, exogenous addition 1.0 mM of a-ketoglutarate delayed germ tube emergence (4 h) compared with the control. On the other hand, in the presence of 1.0 mM glutamate an earlier onset of the germ tube formation was noted. The morphological (monomorphic) mutants, Y-2 and Y-5, showed a high GDH ratio and maintained the yeast morphology.The dimorphic zygomycete Benjaminiella poitrasii has the potential to differentiate into alternative morphologies depending on growth temperature and/or nutritional conditions. The organism grows in mycelial form (M form) at 28°C whereas development of the yeast form (Y form) occurs at 37°C. In addition, optimal yeast growth requires the presence of a hexose as a carbon source as well as an organic amino compound as a nitrogen source. In contrast, the mycelial growth occurs on a variety of carbon sources and with ammonium salts as the sole nitrogen source (5). These observations suggest differential expression of some key enzymes of carbon and nitrogen metabolism in the two morphological forms. Generally, in fungi, the carbon and nitrogen metabolisms are connected via ammonia assimilation and the participating enzymes are NAD-dependent (EC 1.4.1.2) and NADP-dependent (EC 1.4.1.4) glutamate dehydrogenase (GDH), glutamate synthase (GOGAT; EC 1.4.1.13), and glutamine synthetase (GS; EC 6.3.1.2) (8).Since the dimorphic (Y-to-M) transitions in B. poitrasii can be regulated by the carbon-nitrogen balance, the changes in the levels of the aforementioned enzymes were studied to evaluate their possible role. Similar studies were also carried out with monomorphic Y form mutants (Y-2 and Y-5) isolated after N-methyl-N'-nitro-N-nitrosoguanidine treatment of parent strain spores (5). (pH 7.2) was prepared. The pH was adjusted to 7.2 with 2 M NaOH or 1.0 M HCI. The reaction mixture (1 ml) containing 0.4 ml of the above assay mixture, 0.2 ml of crude extract, and 0.35 ml of glass-distilled water was preincubated for 5 min at 37°C. Then the reaction was initiated by the addition MATERUILS AND METHODS

Regulation of chitin synthase activity in the dimorphic fungus Benjaminiella poitrasii by external osmotic pressure

“…The yeast‐mycelium transition experiments were carried out as described earlier [7, 8]. Yeast inoculum was grown in YP5G medium for 24 h at 30°C and the transition was studied in YP medium at 30°C.…”

“…Yeast inoculum was grown in YP5G medium for 24 h at 30°C and the transition was studied in YP medium at 30°C. The morphology (yeast/mycelium) was determined as described earlier[8].…”

Disruption of MRG19 results in altered nitrogen metabolic status and defective pseudohyphal development in Saccharomyces cerevisiae

“…and alters the expression of GDH1 and GDH3, which encode the key enzymes in ammonia assimilation. In Benjaminiella poitrasii, dimorphic transition is correlated with differential expression of different NADP-glutamate dehydrogenases and the carbon/nitrogen balance (Khale et al, 1992). Based on this information, we studied the role of MRG19 in the formation of pseudohyphae.…”

Possible involvement of cyclic adenosine 3',5'-monophosphate in the regulation of NADP-/NAD-glutamate dehydrogenase ratio and in yeast-mycelium transition of Benjaminiella poitrasii

“…The earlier work on the dimorphic behavior of Benjaminiella poitrasii and its morphological mutants (Y-2 and Y-5) has shown that the NADP-/NAD-GDH ratio (GDH ratio) is correlated with the Y*->M transition (7). A (6).…”

“…Transition studies (Y-+M) and determination of morphology were carried out as described earlier (7). Preparation of cell extract and enzyme assays of NAD-and NADP-dependent GDH by reductive amination of a-ketoglutarate as well as protein determination were also carried out as described previously (7). The 3 h when the incubation temperature is changed from 37°C to 28°C.…”