The mineral nutrient needed in greatest abundance by plants is nitrogen. Plants, however, must compete for nitrogen in the soil with abiotic and biotic processes such as erosion, leaching, and microbial consumption. Soil nitrogen is also lost when crops are harvested and plant material is removed from the soil. To be competitive, plants have evolved several mechanisms to acquire nitrogen at low concentrations and to use a variety of forms of nitrogen. Plants can assimilate inorganic forms, such as nitrate and ammonia, and organic forms, such as urea. Some plants, including legumes, can fix dinitrogen gas in association with symbiotic bacteria (see Mylona et al., 1995, this issue). This review focuses on the assimilation of nitrate, a transient yet critical form for plants. Two key questions arise when considering nitrate assimilation. First, how do plants acquire optimal quantities of nitrate when the soil nitrate concentration can vary from 10 pM to 100 mM? Second, how is nitrate assimilation integrated into the overall metabolic program of plants so that optimal amounts of energy and carbon are provided? What is known about nitrate assimilation at the cellular leve1 is summarized in Figure 1. Once taken up, nitrate is either stored in the vacuole or reduced to nitrite by nitrate reductase (NR). Nitrite enters the chloroplast (or plastid in the root) and is then reduced to ammonja by nitrite reductase (NiR). Ammonia can then be fixed into carbon by the action of glutamine synthetase (see Lam et al., 1995, this issue). Reducing energy is provided in the form of NAD(P)H for NR and reduced ferredoxin for NiR. Glutamate provides the immediate carbon skeleton. Because nitrate assimilation produces several hydroxide ions, organic acids are also required for pH homeostasis. Coordinating these steps is a regulatory network that is responsive to both interna1 and external signals, some of which are diagrammed in Figure 1. Ultimately, it is the understanding of this regulatory network that will answer the aforementioned key questions. Genetic analysis of nitrate assimilation has been possible because mutants blocked in the pathway can be rescued by providing ammonia as a source of nitrogen. Such mutants are obtained either by screening directly with an in vitro assay for plants defective in nitrate reduction or by selecting plants that are resistant to chlorate, the chlorine analog of nitrate. When chlorate is taken up and reduced by NR, toxic chlorite is produced. In higher plants, chlorate-resistant mutants are impaired in nitratelchlorate reduction because they have either a defective nitrate reductase structural gene (NIA) or a defective molybdenum cofactor gene (CNX). The one exception is the chll mutant of Arabidopsis, which is defective in nitrate/chlorate uptake. Only in fungi and algae have regulatory mutants been descri bed. There are many excellent recent reviews of the nitrate assimilation field (Pelsy and Caboche, 1992; Warner and Kleinhofs, 1992; Cove, 1993; Crawford and Arst, 1993; Marzluf, 1993; Crawford, 1994; Hoff et al., 1994; Huppe and Turpin, 1994). This review provides an update on recent molecular advances that have uncovered genes and mechanisms responsible for nitrate uptake, reduction, and regulation.

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Nitrate: nutrient and signal for plant growth.

In the fungi, nitrogen metabolism is controlled by a complex genetic regulatory circuit which ensures the preferential use of primary nitrogen sources and also confers the ability to use many different secondary nitrogen sources when appropriate. Most structural genes encoding nitrogen catabolic enzymes are subject to nitrogen catabolite repression, mediated by positive-acting transcription factors of the GATA family of proteins. However, certain GATA family members, such as the yeast DAL80 factor, act negatively to repress gene expression. Selective expression of the genes which encode enzymes for the metabolism of secondary nitrogen sources is often achieved by induction, mediated by pathway-specific factors, many of which have a GAL4-like C6/Zn2 DNA binding domain. Regulation within the nitrogen circuit also involves specific protein-protein interactions, as exemplified by the specific binding of the negative-acting NMR protein with the positive-acting NIT2 protein of Neurospora crassa. Nitrogen metabolic regulation appears to play a significant role in the pathogenicity of certain animal and plant fungal pathogens.

https://mmbr.asm.org/content/mmbr/61/1/17.full.pdf

Genetic regulation of nitrogen metabolism in the fungi.

Metabolic pathway gene clusters in filamentous fungi

Aspergillus fumigatus, a filamentous fungus producing bluish-green conidia, is an important opportunistic pathogen that primarily affects immunocompromised patients. Conidial pigmentation of A. fumigatus significantly influences its virulence in a murine model. In the present study, six genes, forming a gene cluster spanning 19 kb, were identified as involved in conidial pigment biosynthesis in A. fumigatus. Northern blot analyses showed the six genes to be developmentally regulated and expressed during conidiation. The gene products of alb1 (for “albino 1”), arp1 (for “aspergillus reddish-pink 1”), and arp2 have high similarity to polyketide synthases, scytalone dehydratases, and hydroxynaphthalene reductases, respectively, found in the dihydroxynaphthalene (DHN)-melanin pathway of brown and black fungi. The abr1 gene (for “aspergillus brown 1”) encodes a putative protein possessing two signatures of multicopper oxidases. The abr2 gene product has homology to the laccase encoded by the yA gene of Aspergillus nidulans. The function of ayg1 (for “aspergillus yellowish-green 1”) remains unknown. Involvement of the six genes in conidial pigmentation was confirmed by the altered conidial color phenotypes that resulted from disruption of each gene in A. fumigatus. The presence of a DHN-melanin pathway in A. fumigatus was supported by the accumulation of scytalone and flaviolin in the arp1 deletant, whereas only flaviolin was accumulated in the arp2 deletants. Scytalone and flaviolin are well-known signature metabolites of the DHN-melanin pathway. Based on DNA sequence similarity, gene disruption results, and biochemical analyses, we conclude that the 19-kb DNA fragment contains a six-gene cluster which is required for conidial pigment biosynthesis in A. fumigatus. However, the presence of abr1, abr2, and ayg1 in addition to alb1, arp1, and arp2 suggests that conidial pigment biosynthesis in A. fumigatus is more complex than the known DHN-melanin pathway.

/pdf/a-developmentally-regulated-gene-cluster-involved-in-2injpxu1sw.pdf

A developmentally regulated gene cluster involved in conidial pigment biosynthesis in Aspergillus fumigatus.

Nitrogen is the mineral nutrient that plants need in the greatest quantities and the one that most frequently limits plant growth and crop yields. Most plants get their nitrogen (N) from the soil as either nitrate or ammonium, with some species showing a strong preference for one ionic form over the other. The uptake of nitrate and ammonium ions by roots involves a complex set of membrane transport systems that includes both high- and low-affinity transporters; net uptake rates can also be strongly influenced by the rate at which these ions efflux from root cells. Here we review our current picture of the mechanisms responsible for the uptake and efflux of nitrate and ammonium, attempting to integrate the large body of physiological data with the recent advances in the molecular biology of nitrate and ammonium transporters in bacteria and algae as well as in higher plants. We also review what is known at the physiological and molecular levels about the regulation of the N uptake systems, a process which involves both positive signals from soil nitrate or ammonium and feedback inhibitory signals that are generated by the plant's internal N status

M.A.D. Brow

Papers

Isolation and characterisation of the crnA-niiA-niaD gene cluster for nitrate assimilation in Aspergillus nidulans.