Polyamine metabolism in mycorrhizal fungi

Zarb, John (1994) Polyamine metabolism in mycorrhizal fungi. PhD thesis, University of Glasgow.

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Abstract

The polyamine biosynthetic enzymes arginine decarboxylase (ADC), ornithine decarboxylase (ODC), S-adenosylmethionine decarboxylase (AdoMetDC), and spermidine synthase, are susceptible to inhibition by a variety of compounds originally developed for the inhibition of polyamine biosynthesis in animal cells as an approach to cancer treatment. These compounds have subsequently been used in the study of polyamine metabolism in plants, fungi, bacteria, and protozoa, with important consequences. For example, most fungi are considered to possess only ODC for the initial step in polyamine biosynthesis, whilst plants have both ODC and ADC enzymes. Therefore, specific inhibition of ornithine decarboxylation in a plant-pathogenic fungus should inhibit fungal growth without damaging tire host plant. In fact, such an approach to plant disease control has been realised. Difluoromethylarginine (DFMA) and difluoromethylornithine (DFMO) are inhibitors of ADC and ODC, respectively. Cyclohexylamine (CHA) is a spermidine synthase inhibitor, and methylglyoxal bis (guanylhydrazone) (MGBG), an inhibtor of AdoMetDC. Mycorrhizal fungi, which are intimately associated with most plants, and which are of benefit in terrestrial ecosystems, may be vulnerable to compounds affecting polyamine metabolism. This premise was investigated in the ectomycorrhizal fungi Laccaria proximo, Paxillus involutus, Hebeloma mesophaeum, and Thelephora terrestris. Furthermore, in view of the pivotal role of polyamines in organisms, the importance of polyamines in stress responses in plants, and the fact that many mycorrhizal fimgi are tolerant of industrially-polluted soils, polyamine metabolism was studied in P. involutus following exposure of the fungus to toxic metals. Exposure of L. proximo to DFMO led to decreased growth, whilst exposure of the fungus to DFMA enhanced growth. The following data indicate that L. proximo possesses both ADC and ODC activities: (1) DFMA depleted putrescine and did not inhibit ODC activity; (2) incubation of mycelium with a [U-14C]arginine substrate led to [C]putrescine formation; (3) DFMA completely inhibited the formation of polyamines from [U-14C]arginine; (4) DFMO inhibited bound biosynthetic enzyme activity and the formation of putrescine from [U-14C]ornithine. Although exposure of P. involutus to DFMO resulted in reduced growth, and reduced activities of ODC, AdoMetDC and diamine oxidase, this was not accompanied by reductions in polyamine concentrations. Indeed, spermine concentration was substantially increased. Together, these data suggest the existence of a mechanism in P. involutus to maintain intracellular concentrations of free polyamines. H. mesophaeum exhibited a notable lack of response to inhibitor treatments, the possibilities for which are discussed in Chapter 3. The substantial depletion in spermine in T. terrestris, leading to a reduction in growth, following DFMO treatment, suggests tlie importance of this polyamine for growth in T. terrestris, in common with other fungi. Radiolabelled compounds that co-chromatographed with authentic standards of cadaverine, aminopropylcadaverine (APC) and N,N' bis (3 aminopropyl)cadaverine (3 APC), were isolated from reaction mixtures after the decarboxylation of [U-14C]lysine by fungal extracts. The identity of 3 APC was confirmed by nuclear magnetic resonance spectroscopy. In subsequent work, the inhibition of AdoMetDC and spermidine synthase led to significant reductions in the recovery of radiolabelled 3 APC. Results show that a range of ectomycorrhizal and plant-pathogenic fungi can synthesise cadaverine and its higher homologues APC and 3 APC, in reactions catalysed by AdoMetDC and spermidine synthase. Furthermore, these compounds may also be synthesised from the condensation of cadaverine and L-aspartic-p-semialhehyde. Biosynthetic enzyme activity, polyamine concentrations and the rate of conversion of lysine into cadaverine were measured in P. involutus following exposure of the fungus to copper, nickel, lead, or zinc. Metal ions were accumulated in the fungus. Although the concentrations of putrescine, spermidine and spermine were affected to some extent, with copper and nickel treatments associated with reductions in spermidine concentration, these variations in polyamine concentrations did not significantly affect fungal fresh weight. Accumulation of metals in fungal tissues, and exclusion of ions from the cell by sequestration at extracellular sites, are among the mechanisms that would tend to limit metal toxicity in mycorrhizal plants.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Additional Information: Adviser: Dale Walters
Keywords: Microbiology
Date of Award: 1994
Depositing User: Enlighten Team
Unique ID: glathesis:1994-72863
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 11 Jun 2019 11:06
Last Modified: 11 Jun 2019 11:06
URI: http://theses.gla.ac.uk/id/eprint/72863

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