Ribosome Engineering and Secondary Metabolite Production

Ochi, Kozo; Okamoto, Susumu; Tozawa, Yuzuru; Inaoka, Tsukasa; Hosaka, Takeshi; Xu, Jun; Kurosawa, Kazu

doi:10.1016/s0065-2164(04)56005-7

Cited by 140 publications

(119 citation statements)

References 73 publications

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“…Recent X-ray crystallography of the RNA polymerase-ppGpp complex demonstrated unambiguously that there is binding of ppGpp adjacent to the RNA polymerase active center (2). We reported previously that the loss of actinorhodin production observed in relA or relC mutants (which are deficient in ppGpp synthesis) could be suppressed by introducing particular mutations into the rpoB gene that encodes the ␤ subunit of RNA polymerase, which is believed to be a primary target of ppGpp in eliciting changes in gene expression (40,62). Therefore, we assumed that introduction of certain rpoB mutations into KO-350 should restore actinorhodin production to levels that exceed the level in the wild-type strain.…”

Section: Vol 188 2006mentioning

confidence: 95%

EshA Accentuates ppGpp Accumulation and Is Conditionally Required for Antibiotic Production in Streptomyces coelicolor A3(2)

Saito

Hosaka

et al. 2006

J Bacteriol

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Disruption of eshA, which encodes a 52-kDa protein that is produced late during the growth of Streptomyces coelicolor A3(2), resulted in elimination of actinorhodin production. In contrast, disruption of eshB, a close homologue of eshA, had no effect on antibiotic production. The eshA disruptant accumulated lower levels of ppGpp than the wild-type strain accumulated. The loss of actinorhodin production in the eshA disruptant was restored by expression of a truncated relA gene, which increased the ppGpp level to the level in the wild-type strain, indicating that the reduced ppGpp accumulation in the eshA mutant was solely responsible for the loss of antibiotic production. Antibiotic production was also restored in the eshA mutant by introducing mutations into rpoB (encoding the RNA polymerase ␤ subunit) that bypassed the requirement for ppGpp, which is consistent with a role for EshA in modulating ppGpp levels. EshA contains a cyclic nucleotide-binding domain that is essential for its role in triggering actinorhodin production. EshA may provide new insights and opportunities to unravel the molecular signaling events that occur during physiological differentiation in streptomycetes.One of the most intriguing challenges in biology today is elucidation of the mechanisms by which cells detect and respond to extracellular nutritional conditions. Among the prokaryotes, Bacillus subtilis and Streptomyces spp. provide tractable experimental systems for studying such mechanisms because they exhibit a wide range of adaptations to extreme nutrient limitation, including the production and secretion of antibiotics and enzymes and the formation of aerial mycelium (Streptomyces spp.) and endospores (Bacillus spp.) (12). Nutritional status and sporulation have been successfully linked in B. subtilis (45), in which CodY detects and responds to nutrient limitation by monitoring the level of the intracellular GTP pool as an overall indicator of cellular physiology. Recent work by Inaoka and Ochi (20) supported this proposal. The stringent response, a general and ubiquitous response to nutrient limitation in prokaryotes, plays a central role in responding to nutrient stress, mediating its effect through the nucleotide guanosine 5Ј-diphosphate 3Ј-diphosphate (ppGpp) (5). By analyzing mutants with impaired abilities to elicit the stringent response, we have shown that ppGpp plays a role in triggering the onset of antibiotic production in both B. subtilis (21) and Streptomyces spp. (6,7,15,29,37,39,41,52,55), whereas morphological differentiation is triggered by reduced levels of GTP. Streptomycetes are gram-positive, filamentous soil bacteria that have a complex process of morphological differentiation and the ability to produce a wide variety of secondary metabolites (referred to as physiological differentiation) that include antibiotics and other useful medicinal compounds. Morphological differentiation and physiological differentiation in streptomycetes often coincide and occur in response to environmental signals that include nutr...

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Section: Vol 188 2006mentioning

confidence: 95%

EshA Accentuates ppGpp Accumulation and Is Conditionally Required for Antibiotic Production in Streptomyces coelicolor A3(2)

Saito

Hosaka

et al. 2006

J Bacteriol

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“…Upon observing the enhanced production of certain secondary metabolites using the str and rif phenotypes, Ochi and colleagues suspected that ppGpp was involved. However, numerous lines of evidence showed that the stringent response did not play a role (Ochi et al 2004;Ochi 2007). Most importantly, direct measurement of the levels of ppGpp showed that they did not change significantly in the wt and mutant strains.…”

Section: Mechanismmentioning

confidence: 99%

Antibiotic dialogues: induction of silent biosynthetic gene clusters by exogenous small molecules

Okada¹,

Seyedsayamdost²

2016

FEMS Microbiology Reviews

179

150

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One sentence summary: Microbial secondary metabolites represent a significant source of potential drug leads; however, the majority of the corresponding biosynthetic genes are not expressed under normal laboratory conditions. In this review, we assess the capacity of exogenous small molecules, especially antibiotics, to activate these silent gene clusters. Editor: Aimee Shen ABSTRACTNatural products have traditionally served as a dominant source of therapeutic agents. They are produced by dedicated biosynthetic gene clusters that assemble complex, bioactive molecules from simple precursors. Recent genome sequencing efforts coupled with advances in bioinformatics indicate that the majority of biosynthetic gene clusters are not expressed under normal laboratory conditions. Termed 'silent' or 'cryptic', these gene clusters represent a treasure trove for discovery of novel small molecules, their regulatory circuits and their biosynthetic pathways. In this review, we assess the capacity of exogenous small molecules in activating silent secondary metabolite gene clusters. Several approaches that have been developed are presented, including coculture techniques, ribosome engineering, chromatin remodeling and high-throughput elicitor screens. The rationale, applications and mechanisms attendant to each are discussed. Some general conclusions can be drawn from our analysis: exogenous small molecules comprise a productive avenue for the discovery of cryptic metabolites. Specifically, growth-inhibitory molecules, in some cases clinically used antibiotics, serve as effective inducers of silent biosynthetic gene clusters, suggesting that old antibiotics may be used to find new ones. The involvement of natural antibiotics in modulating secondary metabolism at subinhibitory concentrations suggests that they represent part of the microbial vocabulary through which inter-and intraspecies interactions are mediated.

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“…6 Our laboratory has previously developed a method to increase antibacterial production. [7][8][9] This new approach, called 'ribosome engineering' , 10,11 has several advantages. In this method, bacteria are grown on antibiotics to select antibioticresistant strains.…”

Section: Introductionmentioning

confidence: 99%

Rare earth elements activate the secondary metabolite–biosynthetic gene clusters in Streptomyces coelicolor A3(2)

2010

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Genome sequencing projects have revealed many biosynthesis gene clusters for the production of as-yet unknown secondary metabolites, especially in actinomycetes. Here, we report that the rare earth elements, scandium and/or lanthanum, markedly activate, ranging from 2.5-to 12-fold, the expression of nine genes belonging to nine secondary metabolite-biosynthetic gene clusters of Streptomyces coelicolor A3(2) when added to the medium at low concentrations. HPLC analysis of ethyl acetateextractable metabolites indicated the detectability of several compounds only in the rare earth-treated cultures. This approach should facilitate discovery of new biologically active compounds and the study of secondary metabolite production.

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Ribosome Engineering and Secondary Metabolite Production

Cited by 140 publications

References 73 publications

EshA Accentuates ppGpp Accumulation and Is Conditionally Required for Antibiotic Production in Streptomyces coelicolor A3(2)

EshA Accentuates ppGpp Accumulation and Is Conditionally Required for Antibiotic Production in Streptomyces coelicolor A3(2)

Antibiotic dialogues: induction of silent biosynthetic gene clusters by exogenous small molecules

Rare earth elements activate the secondary metabolite–biosynthetic gene clusters in Streptomyces coelicolor A3(2)

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