Abstract NsrR from Streptomyces coelicolor ( Sc) regulates the expression of three genes through the progressive degradation of its [4Fe–4S] cluster on nitric oxide (NO) exposure. We report the 1.95 Å resolution crystal structure of dimeric holo-ScNsrR and show that the cluster is coordinated by the three invariant Cys residues from one monomer and, unexpectedly, Asp8 from the other. A cavity map suggests that NO displaces Asp8 as a cluster ligand and, while D8A and D8C variants remain NO sensitive, DNA binding is affected. A structural comparison of holo-ScNsrR with an apo-IscR-DNA complex shows that the [4Fe–4S] cluster stabilizes a turn between ScNsrR Cys93 and Cys99 properly oriented to interact with the DNA backbone. In addition, an apo ScNsrR structure suggests that Asn97 from this turn, along with Arg12, which forms a salt-bridge with Asp8, are instrumental in modulating the position of the DNA recognition helix region relative to its major groove.
Abstract Streptomyces bacteria are ubiquitous in soils and are well-known for producing secondary metabolites, including antimicrobials. Increasingly, they are being isolated from plant roots and several studies have shown they are specifically recruited to the rhizosphere and the endosphere of the model plant Arabidopsis thaliana . Here we test the hypothesis that Streptomyces bacteria have a beneficial effect on A. thaliana growth and could potentially be used as plant probiotics. To do this, we selectively isolated streptomycetes from surface washed A. thaliana roots and generated high quality genome sequences for five strains which we named L2, M2, M3, N1 and N2. Re-infection of A. thaliana plants with L2, M2 and M3 significantly increased plant biomass individually and in combination whereas N1 and N2 had a negative effect on plant growth, likely due to their production of polyene natural products which can bind to phytosterols and reduce plant growth. N2 exhibits broad spectrum antimicrobial activity and makes filipin-like polyenes, including 14-hydroxyisochainin which inhibits the Take-all fungus, Gaeumannomyces graminis var. tritici . N2 antifungal activity as a whole was upregulated ~2-fold in response to indole-3-acetic acid (IAA) suggesting a possible role during competition in the rhizosphere. Furthermore, coating wheat seeds with N2 spores protected wheat seedlings against Take-all disease. We conclude that at least some soil dwelling streptomycetes confer growth promoting benefits on A. thaliana while others might be exploited to protect crops against disease. Importance It is vital that we reduce our reliance on agrochemicals and there is increasing interest in using bacterial strains to promote plant growth and protect against disease. Our study follows up reports that Arabidopsis thaliana specifically recruits Streptomyces bacteria to its roots. In particular, we test the hypothesis that these bacteria can offer benefits to their A. thaliana hosts and that strains isolated from these plants might be used as probiotics. We isolated Streptomyces strains from surface washed A. thaliana roots and genome sequenced five phylogenetically distinct strains. Genome mining and bioassays indicated that all five strains have plant growth promoting properties, including production of IAA, siderophores and ACC deaminase activity. Three strains significantly increased A. thaliana growth in vitro and when applied in combination in soil. Another produces potent filipin-like antifungal metabolites and we used it as a seed coating to protect germinating wheat seeds against the fungal pathogen Gaeumannomyces graminis var. tritici (wheat Take-all fungus). We conclude that introducing an optimal combination of Streptomyces strains into the root microbiome can provide significant benefits to plants.
The rsmA gene of Streptomyces coelicolor lies directly upstream of the gene encoding the group 3 sigma factor sigma(M). The RsmA protein is a putative member of the HATPase_c family of anti-sigma factors but is unusual in that it contains seven cysteine residues. Bacterial two-hybrid studies demonstrate that it interacts specifically with sigma(M), and in vitro studies of the purified proteins by native PAGE and transcription assays confirmed that they form a complex. Characterization of RsmA revealed that it binds ATP and that, as isolated, it contains significant quantities of iron and inorganic sulfide, in equal proportion, with spectroscopic properties characteristic of a [2Fe-2S] cluster-containing protein. Importantly, the interaction between RsmA and sigma(M) is dependent on the presence of the iron-sulfur cluster. We propose a model in which RsmA regulates the activity of sigma(M). Loss of the cluster, in response to an as yet unidentified signal, activates sigma(M) by abolishing its interaction with the anti-sigma factor. This represents a major extension of the functional diversity of iron-sulfur cluster proteins.
Significance The evolutionary significance of assortative mating by diet, mediated by gut bacteria is a puzzle, but it has had a huge impact and has provided a keystone to support increasing interest in the “holobiome.” However, in species such as Drosophila melanogaster that have flexible gut microbiomes, any reproductive isolation mediated by gut bacteria specific to host diets can only be transient. Here, we replicated and extended tests of this idea. Despite differences in gut microbiomes, we failed to recover previously observed patterns of nonrandom mating and found no evidence that mating preferences were associated with diet or gut bacteria. This suggests that the evolutionary importance of gut microbiomes in host divergence needs careful consideration on a case-by-case basis.
Abstract Streptomyces bacteria make numerous secondary metabolites, including half of all known antibiotics. Understanding the global regulation of secondary metabolism is important because most Streptomyces natural products are not made under laboratory conditions and unlocking ‘cryptic’ biosynthetic gene clusters (BGCs) is a major focus for natural product discovery. Production is coordinated with sporulation but the regulators that coordinate development with antibiotic biosynthesis are largely unknown. Here we characterise a highly conserved actinobacterial response regulator called MtrA in antibiotic-producing Streptomyces species. We show that MtrA is an essential global regulator of secondary metabolism that directly activates antibiotic production in in S. coelicolor and S. venezuelae . MtrA also controls key developmental genes required for DNA replication and cell division and we propose that MtrA is the missing link that coordinates secondary metabolism with development in Streptomyces species.
Arabidopsis thaliana has a diverse but consistent root microbiome, recruited in part by the release of fixed carbon in root exudates. Here we focussed on the recruitment of Streptomyces bacteria, which are well established plant-growth-promoting rhizobacteria and which have been proposed to be recruited to A. thaliana roots by the release of salicylic acid. We generated high quality genome sequences for eight Streptomyces endophyte strains and showed that although some strains do enhance plant growth, they are not attracted to, and do not feed on, salicyclic acid. We used 13CO2 DNA-stable isotope probing to determine which bacteria are fed by the plants in the rhizo- and endosphere and found that streptomycetes did not feed on root exudates in vivo, despite the fact that they can use exudate as sole carbon and nitrogen sources in vitro. We confirmed increased root colonisation by streptomycetes in plants that constitutively produce salicylic acid, but these plants exhibited a pleiotropic phenotype of early senescence and weak growth. We propose that streptomycetes are attracted to the rhizosphere by root exudates but can be outcompeted for this food source by more abundant proteobacteria and most likely feed off unlabelled complex organic matter.
Antimycins are an extended family of depsipeptides that are made by filamentous actinomycete bacteria and were first isolated more than 60 years ago. Recently, antimycins have attracted renewed interest because of their activities against the anti-apoptotic machineries inside human cells which could make them promising anti-cancer compounds. The biosynthetic pathway for antimycins was recently characterised but very little is known about the organisation and regulation of the antimycin (ant) gene cluster. Here we report that the ant gene cluster in Streptomyces albus is organized into four transcriptional units; the antBA, antCDE, antGF and antHIJKLMNO operons. Unusually for secondary metabolite clusters, the antG and antH promoters are regulated by an extracytoplasmic function (ECF) RNA polymerase sigma factor named σAntA which represents a new sub-family of ECF σ factors that is only found in antimycin producing strains. We show that σAntA controls production of the unusual precursor 3-aminosalicylate which is absolutely required for the production of antimycins. σAntA is highly conserved in antimycin producing strains and the −10 and −35 elements at the σAntA regulated antG and antH promoters are also highly conserved suggesting a common mechanism of regulation. We also demonstrate that altering the C-terminal Ala-Ala residues found in all σAntA proteins to Asp-Asp increases expression of the antFG and antGHIJKLMNO operons and we speculate that this Ala-Ala motif may be a signal for the protease ClpXP.
The high critical magnetic field and relatively high critical temperature of niobium nitride (NbN) make it a promising material candidate for applications in superconducting quantum technology. However, NbN-based devices and circuits are sensitive to decoherence sources such as two-level system (TLS) defects. Here, we numerically and experimentally investigate NbN superconducting microwave coplanar waveguide resonator arrays, with a 100 nm thickness, capacitively coupled to a common coplanar waveguide on a silicon chip. We observe that the resonators' internal quality factor (Qi) decreases from Qi ~ 1.07*10^6 in a high power regime (< nph > = 27000) to Qi ~ 1.36 *10^5 in single photon regime at temperature T = 100 mK. Data from this study is consistent with the TLS theory, which describes the TLS interactions in resonator substrates and interfaces. Moreover, we study the temperature dependence internal quality factor and frequency tuning of the coplanar waveguide resonators to characterise the quasiparticle density of NbN. We observe that the increase in kinetic inductance at higher temperatures is the main reason for the frequency shift. Finally, we measure the resonators' resonance frequency and internal quality factor at single photon regime in response to in-plane magnetic fields B||. We verify that Qi stays well above 10^4 up to B|| = 240 mT in the photon number < nph > = 1.8 at T = 100 mK. Our results may pave the way for realising robust microwave superconducting circuits for circuit quantum electrodynamics (cQED) at high magnetic fields necessary for fault-tolerant quantum computing, and ultrasensitive quantum sensing.
Many, if not all, plants and animals form mutually beneficial symbioses (mutualisms) with microbes and a subset of these mutualisms are defensive, in which the host provides food and housing in return for defence against disease. These symbioses typically involve antibiotic-producing bacteria, the best known of which are filamentous actinomycetes in the genera Streptomyces and Pseudonocardia and unicellular species in the genus Pseudomonas. Such mutualisms are likely to be widespread in nature, but they are best characterised in insects, which provide experimentally tractable models for studying symbiosis and microbiome formation because they typically host less complex microbial communities. Here, we examine the mutualisms formed between insects and antibiotic-producing bacteria using well-characterised examples, including digger wasps and their endosymbiotic Streptomyces species, attine ants and their mutualist Pseudonocardia species and Paederus beetles with their pederin-producing Pseudomonas species. We also discuss how searching such symbiotic niches can give insights into the evolution and functions of microbial specialised metabolites and provide new platforms for antibiotic discovery.
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