Nitric oxide synthase inhibitors and nitric oxide donors modulate the biosynthesis of thaxtomin A, a nitrated phytotoxin produced by Streptomyces spp.

doi:10.1016/j.niox.2004.11.004

Nitric Oxide

Volume 12, Issue 1, February 2005, Pages 46-53

https://doi.org/10.1016/j.niox.2004.11.004 Get rights and content

Abstract

Evidence for the involvement of a bacterial nitric oxide synthase (NOS) in the biosynthesis of a phytotoxin is presented. Several species of Streptomyces bacteria produce secondary metabolites with unusual nitrogen groups, such as thaxtomin A (ThxA), which contains a nitroindole moiety. ThxA is a phytotoxin made by three pathogenic Streptomyces species that cause common scab of potato. All three species possess a gene homologous to the oxygenase domain of murine inducible NOS, and this gene, nos, is essential for normal levels of ThxA production. We grew Streptomyces turgidiscabies in the presence of several known NOS inhibitors and a nitric oxide (NO) scavenger to determine their effect on ThxA production. The NO scavenger (CPTIO) and four NOS inhibitors (NAME, NMMA, AG, and 7-NI) reduced ThxA production without affecting bacterial growth. A strain of S. turgidiscabies from which the nos gene had been deleted was grown in the presence of three NO donors (DEANO, SIN, and SNAP), and all three partially restored ThxA production. Our data suggest that bacterial nitric oxide synthases may, at least in part, produce NO for biosynthetic purposes, rather than for cellular signaling, as they do in mammals.

Section snippets

Materials

Chlorzoxazone was obtained from Acros Organics-Fischer (Pittsburgh, Pennsylvania). Miconazole and 3-morpholinosydnonimine were obtained from Sigma–Aldrich (St. Louis, Missouri). 2-(4-Carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide and diethylamino nitric oxide were obtained from Molecular Probes (Eugene, Oregon). All other chemicals were from Calbiochem (La Jolla, CA). Oat bran was obtained from New Hope Mills (Moravia, New York).

Bacterial strains

Assays were performed with S. turgidiscabies

Effect of NMMA on ThxA production

l-NMMA, an arginine analog and a NOS inhibitor, has been studied extensively and is known to be a non-selective inhibitor of all human NOS isoforms [2], [8]. To determine if the catalytic activity of stNOS was affected by a NOS inhibitor, we grew S. turgidiscabies in the presence of l-NMMA, reasoning that if stNOS had any structural and functional similarities to mammalian NOSs, l-NMMA might interact with the enzyme. We found that all concentrations of NMMA tested reduced ThxA production, and

Discussion

Three Streptomyces species which cause common scab of potato possess nos, a gene encoding a nitric oxide synthase. Although stNOS is highly homologous to the oxygenase domain of several mammalian NOSs, the data presented here indicate that the function of stNOS is necessary for the formation of nitro-tryptophan as an essential component of a secondary metabolic pathway for phytotoxin biosynthesis. We have shown that NOS inhibitors, NAME, NMMA, AG, and 7-NI, as well as the NO scavenger CPTIO,

Acknowledgments

We thank Brian Crane for helpful discussions in the course of this study. This work was supported by the Cooperative State Research, Education, and Extension Service, US Department of Agriculture, under Agreement No. 2002-30001-12096. Mention of a trademark, proprietary product, or vendor does not constitute a guarantee or warranty of the product by the US Department of Agriculture and does not imply its approval to the exclusion of other vendors that may be suitable.

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However, most bacterial NOSs show similar structural, spectroscopic, catalytic properties to mammalian enzyme with few interesting exceptions (Gusarov et al., 2008). The functions of NOS in bacteria vary considerably from mammals ranging from biosynthesis of phytotoxin called thaxtomin by site specific nitration in Streptomyces turgidiscabies (Wach et al., 2005), resistance to oxidative stress in Bacillus subtilis (Gusarov et al., 2005), recovery from UV-B radiation damage in Deinococcus radiodurans and in regulating the transition from aerobic to nitrate dependent respiration during microaerobiasis in Staphylococcus aureus (Patel et al., 2009; Mogen et al., 2017). In photosynthetic organism, the first NOS was characterised in Ostreococcus tauri, a marine unicellular algae having identical characterstics to animal NOS.
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Thaxtomin phytotoxins, first reported in 1989, are cyclic dipeptides (2,5-diketopiperazines) formed from the condensation of 4-nitrotrytophan and phenylalanine groups. Inclusion of a 4-nitroindole moiety that is an essential requirement for their phytotoxicity makes them unique amongst microbial generated metabolites. Individual thaxtomins differ only in the presence or absence of N-methyl and hydroxyl groups and their respective substitution sites. The name “thaxtomin” was assigned to these compounds in honor of the eminent American phytopathologist Roland Thaxter who first described the nature of the microorganisms responsible for their production. The great interest in the thaxtomins derives mainly from their established roles as virulence factors in the common scab of potato disease and their apparent ability to inhibit cellulose synthesis in developing plant cells. Surveys of comparable scab-inducing organisms from potato producing regions throughout the world have confirmed the generality of the phytotoxin associated process. Descriptions of the isolation, chemical structure determination, total synthesis, biosynthesis, transformation by microorganisms and biological activity of the thaxtomins are comprehensively reviewed.
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Nitric oxide synthase inhibitors and nitric oxide donors modulate the biosynthesis of thaxtomin A, a nitrated phytotoxin produced by Streptomyces spp.

Abstract

Section snippets

Materials

Bacterial strains

Effect of NMMA on ThxA production

Discussion

Acknowledgments

Structure

Nitric Oxide

Biochim. Biophys. Acta

Physiol. Mol. Plant Pathol.

J. Biol. Chem.

Curr. Opin. Chem. Biol.

Phytochemistry

Physiol. Mol. Plant Pathol.

J. Biol. Chem.

Biochim. Biophys. Acta

Cloning, expression, and characterization of a nitric oxide synthase protein from Deinococcus radiodurans

Proc. Natl. Acad. Sci. USA

Nitric oxide synthases: structure, function and inhibition

Biochem. J.

Chemical and kinetic study on stabilities of 3-morpholinosydnonimine and its N-ethoxycarbonyl derivative

Chem. Pharm. Bull.

Production and regulation of potato-scab-inducing phytotoxins by Streptomyces scabies

J. Gen. Microbiol.

Complete genome sequence of the model actinomycete Streptomyces coelicolor A3(2)

Nature

Nitric oxide biosynthesis, nitric oxide synthase inhibitors and arginase competition for l-arginine utilization

Cell. Mol. Life Sci.

LL-F42248a, a novel chlorinated pyrrole antibiotic

J. Antibiot.

Direct biochemical nitration in the biosynthesis of dioxapyrrolomycin. A unique mechanism for the introduction of nitro groups in microbial products,

J. Chem. Soc., Chem. Commun.

Actinomycete development, antibiotic production, and phylogeny: questions and challenges

Purification and characterization of nitric oxide synthase (NOSNoc) from a Nocardia species

J. Bacteriol.

Nitric oxide induces the apoptosis of human BCR-ABL-positive myeloid leukemia cells: evidence for the chelation of intracellular iron

Leukemia

Regulation of nitric oxide synthase expression and activity

Purification and characterization of nitric oxide synthase (NOS_Noc) from a Nocardia species