Publications by authors named "Paula Yagüe"

21 Publications

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Correction: Yagüe, P., et al. Goals and Challenges in Bacterial Phosphoproteomics. 2019, , 5678.

Int J Mol Sci 2020 Dec 9;21(24). Epub 2020 Dec 9.

Área de Microbiología, Departamento de Biología Funcional, IUOPA, ISPA, Facultad de Medicina, Universidad de Oviedo, 33006 Oviedo, Spain.

The authors wish to make the following corrections to this paper [1]:The author name "Gemma Fernánez-García" should be "Gemma Fernández-García" [...].
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http://dx.doi.org/10.3390/ijms21249381DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7764465PMC
December 2020

Goals and Challenges in Bacterial Phosphoproteomics.

Int J Mol Sci 2019 Nov 13;20(22). Epub 2019 Nov 13.

Área de Microbiología, Departamento de Biología Funcional, IUOPA, ISPA, Facultad de Medicina, Universidad de Oviedo, 33006 Oviedo, Spain.

Reversible protein phosphorylation at serine, threonine and tyrosine is a well-known dynamic post-translational modification with stunning regulatory and signalling functions in eukaryotes. Shotgun phosphoproteomic analyses revealed that this post-translational modification is dramatically lower in bacteria than in eukaryotes. However, Ser/Thr/Tyr phosphorylation is present in all analysed bacteria (24 eubacteria and 1 archaea). It affects central processes, such as primary and secondary metabolism development, sporulation, pathogenicity, virulence or antibiotic resistance. Twenty-nine phosphoprotein orthologues were systematically identified in bacteria: ribosomal proteins, enzymes from glycolysis and gluconeogenesis, elongation factors, cell division proteins, RNA polymerases, ATP synthases and enzymes from the citrate cycle. While Ser/Thr/Tyr phosphorylation exists in bacteria, there is a consensus that histidine phosphorylation is the most abundant protein phosphorylation in prokaryotes. Unfortunately, histidine shotgun phosphorproteomics is not possible due to the reduced phosphohistidine half-life under the acidic pH conditions used in standard LC-MS/MS analysis. However, considering the fast and continuous advances in LC-MS/MS-based phosphoproteomic methodologies, it is expected that further innovations will allow for the study of His phosphoproteomes and a better coverage of bacterial phosphoproteomes. The characterisation of the biological role of bacterial Ser/Thr/Tyr and His phosphorylations might revolutionise our understanding of prokaryotic physiology.
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http://dx.doi.org/10.3390/ijms20225678DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6888350PMC
November 2019

Nonpathogenic Bacteria as Targets in Antimicrobial High-Throughput Screening.

Trends Microbiol 2020 01 6;28(1):1-2. Epub 2019 Nov 6.

Área de Microbiología, Departamento de Biología Funcional, IUOPA, ISPA, Facultad de Medicina, Universidad de Oviedo, 33006 Oviedo, Spain. Electronic address:

Antimicrobial screening usually analyses the effects of natural or synthetic molecules against pathogens. McAuley et al. changed this paradigm, testing the effect of synthetic compounds against the sporulation of the nonpathogenic bacterium Streptomyces venezuelae. They discovered a novel DNA-targeting antibiotic effective against pathogens.
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http://dx.doi.org/10.1016/j.tim.2019.10.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6978137PMC
January 2020

Cytosolic copper is a major modulator of germination, development and secondary metabolism in Streptomyces coelicolor.

Sci Rep 2019 03 12;9(1):4214. Epub 2019 Mar 12.

Área de Microbiología, Departamento de Biología Funcional, IUOPA and ISPA, Facultad de Medicina, Universidad de Oviedo, 33006, Oviedo, Spain.

Streptomycetes are important biotechnological bacteria with complex differentiation. Copper is a well-known positive regulator of differentiation and antibiotic production. However, the specific mechanisms buffering cytosolic copper and the biochemical pathways modulated by copper remain poorly understood. Here, we developed a new methodology to quantify cytosolic copper in single spores which allowed us to propose that cytosolic copper modulates asynchrony of germination. We also characterised the SCO2730/2731 copper chaperone/P-type ATPase export system. A Streptomyces coelicolor strain mutated in SCO2730/2731 shows an important delay in germination, growth and sporulation. Secondary metabolism is heavily enhanced in the mutant which is activating the production of some specific secondary metabolites during its whole developmental cycle, including germination, the exponential growth phase and the stationary stage. Forty per cent of the S. coelicolor secondary metabolite pathways, are activated in the mutant, including several predicted pathways never observed in the lab (cryptic pathways). Cytosolic copper is precisely regulated and has a pleiotropic effect in gene expression. The only way that we know to achieve the optimal concentration for secondary metabolism activation, is the mutagenesis of SCO2730/2731. The SCO2730/2731 genes are highly conserved. Their inactivation in industrial streptomycetes may contribute to enhance bioactive compound discovery and production.
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http://dx.doi.org/10.1038/s41598-019-40876-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6414726PMC
March 2019

Quantitative Proteome and Phosphoproteome Analyses of Reveal Proteins and Phosphoproteins Modulating Differentiation and Secondary Metabolism.

Mol Cell Proteomics 2018 08 21;17(8):1591-1611. Epub 2018 May 21.

From the ‡Área de Microbiología, Departamento de Biología Funcional e IUOPA, Facultad de Medicina, Universidad de Oviedo, 33006 Oviedo, Spain;

Streptomycetes are multicellular bacteria with complex developmental cycles. They are of biotechnological importance as they produce most bioactive compounds used in biomedicine, antibiotic, antitumoral and immunosupressor compounds. genomes encode many Ser/Thr/Tyr kinases, making this genus an outstanding model for the study of bacterial protein phosphorylation events. We used mass spectrometry based quantitative proteomics and phosphoproteomics to characterize bacterial differentiation and activation of secondary metabolism of We identified and quantified 3461 proteins corresponding to 44.3% of the proteome across three developmental stages: vegetative hypha (first mycelium); secondary metabolite producing hyphae (second mycelium); and sporulating hyphae. A total of 1350 proteins exhibited more than 2-fold expression changes during the bacterial differentiation process. These proteins include 136 regulators (transcriptional regulators, transducers, Ser/Thr/Tyr kinases, signaling proteins), as well as 542 putative proteins with no clear homology to known proteins which are likely to play a role in differentiation and secondary metabolism. Phosphoproteomics revealed 85 unique protein phosphorylation sites, 58 of them differentially phosphorylated during differentiation. Computational analysis suggested that these regulated protein phosphorylation events are implicated in important cellular processes, including cell division, differentiation, regulation of secondary metabolism, transcription, protein synthesis, protein folding and stress responses. We discovered a novel regulated phosphorylation site in the key bacterial cell division protein FtsZ (pSer319) that modulates sporulation and regulates actinorhodin antibiotic production. We conclude that manipulation of distinct protein phosphorylation events may improve secondary metabolite production in industrial streptomycetes, including the activation of cryptic pathways during the screening for new secondary metabolites from streptomycetes.
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http://dx.doi.org/10.1074/mcp.RA117.000515DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6072539PMC
August 2018

Streptomyces Differentiation in Liquid Cultures as a Trigger of Secondary Metabolism.

Antibiotics (Basel) 2018 May 14;7(2). Epub 2018 May 14.

Área de Microbiología, Departamento de Biología Funcional IUOPA, Facultad de Medicina, Universidad de Oviedo, 33006 Oviedo, Spain.

is a diverse group of gram-positive microorganisms characterised by a complex developmental cycle. produce a number of antibiotics and other bioactive compounds used in the clinic. Most screening campaigns looking for new bioactive molecules from actinomycetes have been performed empirically, e.g., without considering whether the bacteria are growing under the best developmental conditions for secondary metabolite production. These screening campaigns were extremely productive and discovered a number of new bioactive compounds during the so-called "golden age of antibiotics" (until the 1980s). However, at present, there is a worrying bottleneck in drug discovery, and new experimental approaches are needed to improve the screening of natural actinomycetes. are still the most important natural source of antibiotics and other bioactive compounds. They harbour many cryptic secondary metabolite pathways not expressed under classical laboratory cultures. Here, we review the new strategies that are being explored to overcome current challenges in drug discovery. In particular, we focus on those aimed at improving the differentiation of the antibiotic-producing mycelium stage in the laboratory.
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http://dx.doi.org/10.3390/antibiotics7020041DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6022995PMC
May 2018

ArgR of Is a Pleiotropic Transcriptional Regulator: Effect on the Transcriptome, Antibiotic Production, and Differentiation in Liquid Cultures.

Front Microbiol 2018 1;9:361. Epub 2018 Mar 1.

Área de Microbiología, Departamento de Biología Molecular, Universidad de León, León, Spain.

ArgR is a well-characterized transcriptional repressor controlling the expression of arginine and pyrimidine biosynthetic genes in bacteria. In this work, the biological role of ArgR was analyzed by comparing the transcriptomes of Δ and its parental strain, M145, at five different times over a 66-h period. The effect of ArgR was more widespread than that of the orthologous protein of , affecting the expression of 1544 genes along the microarray time series. This regulator repressed the expression of arginine and pyrimidine biosynthetic genes, but it also modulated the expression of genes not previously described to be regulated by ArgR: genes involved in nitrogen metabolism and nitrate utilization; the , and genes for antibiotic production; genes for the synthesis of the osmotic stress protector ectoine; genes related to hydrophobic cover formation and sporulation (chaplins, rodlins, , and genes); all the genes encoding proteins for glycan cell wall biosynthesis; and genes involved in gas vesicle formation. Many of these genes contain ARG boxes for ArgR binding. ArgR binding to seven new ARG boxes, located upstream or near the , and genes, was tested by DNA band-shift assays. These data and those of previously assayed fragments permitted the construction of an improved model of the ArgR binding site. Interestingly, the overexpression of sporulation genes observed in the Δ mutant in our culture conditions correlated with a sporulation-like process, an uncommon phenotype.
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http://dx.doi.org/10.3389/fmicb.2018.00361DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5839063PMC
March 2018

The SCO4117 ECF Sigma Factor Pleiotropically Controls Secondary Metabolism and Morphogenesis in .

Front Microbiol 2018 21;9:312. Epub 2018 Feb 21.

Área de Microbiología, Departamento de Biología Funcional e IUOPA, Facultad de Medicina, Universidad de Oviedo, Oviedo, Spain.

Extracytoplasmic function (ECF) sigma factors are a major type of bacterial signal-transducers whose biological functions remain poorly characterized in streptomycetes. In this work we studied SCO4117, a conserved ECF sigma factor from the ECF52 family overexpressed during substrate and aerial mycelium stages. The ECF52 sigma factors harbor, in addition to the ECF sigma factor domain, a zinc finger domain, a transmembrane region, a proline-rich C-terminal extension, and a carbohydrate-binding domain. This class of ECF sigma factors is exclusive to Actinobacteria. We demonstrate that is an activator of secondary metabolism, aerial mycelium differentiation, and sporulation, in all the culture media (sucrose-free R5A, GYM, MM, and SFM) analyzed. Aerial mycelium formation and sporulation are delayed in a knockout strain. Actinorhodin production is delayed and calcium-dependent antibiotic production is diminished, in the Δ mutant. By contast, undecylprodigiosin production do not show significant variations. The expression of genes encoding secondary metabolism pathways (deoxysugar synthases, actinorhodin biosynthetic genes) and genes involved in differentiation () was dramatically reduced (up to 300-fold) in the knockout. A putative motif bound, with the consensus "CSGYN-17bps-SRHA" sequence, was identified in the promoter region of 29 genes showing affected transcription in the mutant, including one of the promoters. is a conserved gene with complex regulation at the transcriptional and post-translational levels and the first member of the ECF52 family characterized.
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http://dx.doi.org/10.3389/fmicb.2018.00312DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5826349PMC
February 2018

Biosynthesis of Apigenin, Luteolin, and Eriodictyol in the Actinomycete and Production Improvement by Feeding and Spore Conditioning.

Front Microbiol 2017 30;8:921. Epub 2017 May 30.

Biotechnology in Nutraceuticals and Bioactive Compounds-BIONUC, Departamento de Biología Funcional, Área de Microbiología, University of OviedoOviedo, Spain.

Nutraceutical compounds as plant flavonoids play an important role in prevention and modulation of diverse heath conditions, as they exert interesting antifungal, antibacterial, antioxidant, and antitumor effects. They also possess anti-inflammatory activities in arthritis, cardiovascular disease or neurological diseases, as well as modulatory effects on the CYP450 activity on diverse drugs. Most flavonoids are bioactive molecules of plant origin, but their industrial production is sometimes hindered due to reasons as low concentration in the plant tissues, presence in only some species or as a complex mixture or inactive glycosides in plant vacuolae. In this work, we describe the biosynthesis of two important flavones, apigenin and luteolin, and one known flavanone, eriodictyol. Their plant biosynthetic pathways have been reconstructed for heterologous expression in , an actinomycete bacterium manageable at industrial production level. Also, production levels for apigenin have been improved by feeding with naringenin precursor, and timing for settlement of secondary metabolism has been advanced by spore conditioning. In the cases of eriodictyol and luteolin, their production in this important type of biotechnology-prone bacteria, the actinomycetes, had not been described in the literature yet.
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http://dx.doi.org/10.3389/fmicb.2017.00921DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5447737PMC
May 2017

Subcompartmentalization by cross-membranes during early growth of Streptomyces hyphae.

Nat Commun 2016 08 12;7:12467. Epub 2016 Aug 12.

Área de Microbiología, Departamento de Biología Funcional e IUOPA, Facultad de Medicina, Universidad de Oviedo, 33006 Oviedo, Spain.

Bacteria of the genus Streptomyces are a model system for bacterial multicellularity. Their mycelial life style involves the formation of long multinucleated hyphae during vegetative growth, with occasional cross-walls separating long compartments. Reproduction occurs by specialized aerial hyphae, which differentiate into chains of uninucleoid spores. While the tubulin-like FtsZ protein is required for the formation of all peptidoglycan-based septa in Streptomyces, canonical divisome-dependent cell division only occurs during sporulation. Here we report extensive subcompartmentalization in young vegetative hyphae of Streptomyces coelicolor, whereby 1 μm compartments are formed by nucleic acid stain-impermeable barriers. These barriers possess the permeability properties of membranes and at least some of them are cross-membranes without detectable peptidoglycan. Z-ladders form during the early growth, but cross-membrane formation does not depend on FtsZ. Thus, a new level of hyphal organization is presented involving unprecedented high-frequency compartmentalization, which changes the old dogma that Streptomyces vegetative hyphae have scarce compartmentalization.
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http://dx.doi.org/10.1038/ncomms12467DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4990651PMC
August 2016

Characterization of SCO4439, a D-alanyl-D-alanine carboxypeptidase involved in spore cell wall maturation, resistance, and germination in Streptomyces coelicolor.

Sci Rep 2016 Feb 12;6:21659. Epub 2016 Feb 12.

Área de Microbiología, Departamento de Biología Funcional and IUOPA, Facultad de Medicina, Universidad de Oviedo, 33006 Oviedo, Spain.

This work contributes to the understanding of cell wall modifications during sporulation and germination in Streptomyces by assessing the biological function and biochemical properties of SCO4439, a D-alanyl-D-alanine carboxypeptidase (DD-CPase) constitutively expressed during development. SCO4439 harbors a DD-CPase domain and a putative transcriptional regulator domain, separated by a putative transmembrane region. The recombinant protein shows that DD-CPase activity is inhibited by penicillin G. The spores of the SCO4439::Tn5062 mutant are affected in their resistance to heat and acid and showed a dramatic increase in swelling during germination. The mycelium of the SCO4439::Tn5062 mutant is more sensitive to glycopeptide antibiotics (vancomycin and teicoplanin). The DD-CPase domain and the hydrophobic transmembrane region are highly conserved in Streptomyces, and both are essential for complementing the wild type phenotypes in the mutant. A model for the biological mechanism behind the observed phenotypes is proposed, in which SCO4439 DD-CPase releases D-Ala from peptidoglycan (PG) precursors, thereby reducing the substrate pool for PG crosslinking (transpeptidation). PG crosslinking regulates spore physical resistance and germination, and modulates mycelium resistance to glycopeptides. This study is the first demonstration of the role of a DD-CPase in the maturation of the spore cell wall.
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http://dx.doi.org/10.1038/srep21659DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4751497PMC
February 2016

New ΦBT1 site-specific integrative vectors with neutral phenotype in Streptomyces.

Appl Microbiol Biotechnol 2016 Mar 13;100(6):2797-808. Epub 2016 Jan 13.

Área de Microbiología, Departamento de Biología Funcional e IUOPA, Facultad de Medicina, Universidad de Oviedo, 33006, Oviedo, Spain.

Integrative plasmids are one of the best options to introduce genes in low copy and in a stable form into bacteria. The ΦC31-derived plasmids constitute the most common integrative vectors used in Streptomyces. They integrate at different positions (attB and pseudo-attB sites) generating different mutations. The less common ΦBT1-derived vectors integrate at the unique attB site localized in the SCO4848 gene (S. coelicolor genome) or their orthologues in other streptomycetes. This work demonstrates that disruption of SCO4848 generates a delay in spore germination. SCO4848 is co-transcribed with SCO4849, and the spore germination phenotype is complemented by SCO4849. Plasmids pNG1-4 were created by modifying the ΦBT1 integrative vector pMS82 by introducing a copy of SCO4849 under the control of the promoter region of SCO4848. pNG2 and pNG4 also included a copy of the P ermE * in order to facilitate gene overexpression. pNG3 and pNG4 harboured a copy of the bla gene (ampicillin resistance) to facilitate selection in E. coli. pNG1-4 are the only integrative vectors designed to produce a neutral phenotype when they are integrated into the Streptomyces genome. The experimental approach developed in this work can be applied to create phenotypically neutral integrative plasmids in other bacteria.
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http://dx.doi.org/10.1007/s00253-015-7271-0DOI Listing
March 2016

Transcriptional response to vancomycin in a highly vancomycin-resistant Streptomyces coelicolor mutant.

Future Microbiol 2014 ;9(5):603-22

Instituto de Biotecnología de León (INBIOTEC), Avda. Real 1, 24006 León, Spain.

Aim: The main objective of this study is to understand the mechanism of vancomycin resistance in a Streptomyces coelicolor disrupted mutant highly resistant to vancomycin.

Materials & Methods: Different techniques have been performed in the study including gene disruption, primer extension, antibiotic susceptibility tests, electron microscopy, confocal microscopy, cell wall analysis and microarrays.

Results: During the phenotypical characterization of mutant strains affected in phosphate-regulated genes of unknown function, we found that the S. coelicolor SCO2594 disrupted mutant was highly resistant to vancomycin and had other phenotypic alterations such as antibiotic overproduction, impaired growth and reduction of phosphate cell wall content. Transcriptomic studies with this mutant indicated a relationship between vancomycin resistance and cell wall stress.

Conclusion: We identified a S. coelicolor mutant highly resistant to vancomycin in both high and low phosphate media. In addition to Van proteins, others such as WhiB or SigE appear to be involved in this regulatory mechanism.
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http://dx.doi.org/10.2217/fmb.14.21DOI Listing
April 2015

Cell immobilization of Streptomyces coelicolor : effect on differentiation and actinorhodin production.

Int Microbiol 2014 Jun;17(2):75-80

Microbiology Section, Department of Functional Biology and IUOPA, School of Medicine, University of Oviedo, Spain.

Streptomycetes are mycelium-forming bacteria that produce two thirds of the clinically relevant secondary metabolites. Despite the fact that secondary metabolite production is activated at specific developmental stages of the Streptomyces spp. life cycle, different streptomycetes show different behaviors, and fermentation conditions need to be optimized for each specific strain and secondary metabolite. Cell-encapsulation constitutes an interesting alternative to classical fermentations, which was demonstrated to be useful in Streptomyces, but development under these conditions remained unexplored. In this work, the influence of cell-encapsulation in hyphae differentiation and actinorhodin production was explored in the model Streptomyces coelicolor strain. Encapsulation led to a delay in growth and to a reduction of mycelium density and cell death. The high proportion of viable hyphae duplicated extracellular actinorhodin production in the encapsulated cultures with respect to the non-encapsulated ones.
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http://dx.doi.org/10.2436/20.1501.01.209DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4597334PMC
June 2014

Transcriptomic analysis of liquid non-sporulating Streptomyces coelicolor cultures demonstrates the existence of a complex differentiation comparable to that occurring in solid sporulating cultures.

PLoS One 2014 21;9(1):e86296. Epub 2014 Jan 21.

Área de Microbiología, Departamento de Biología Funcional and IUOPA, Facultad de Medicina, Universidad de Oviedo, Oviedo, Spain.

Streptomyces species produce many clinically relevant secondary metabolites and exhibit a complex development that includes hyphal differentiation and sporulation in solid cultures. Industrial fermentations are usually performed in liquid cultures, conditions in which Streptomyces strains generally do not sporulate, and it was traditionally assumed that no differentiation took place. The aim of this work was to compare the transcriptomes of S. coelicolor growing in liquid and solid cultures, deepening the knowledge of Streptomyces differentiation. Microarrays demonstrated that gene expression in liquid and solid cultures were comparable and data indicated that physiological differentiation was similar for both conditions. Eighty-six percent of all transcripts showed similar abundances in liquid and solid cultures, such as those involved in the biosynthesis of actinorhodin (actVA, actII-4) and undecylprodigiosin (redF); activation of secondary metabolism (absR1, ndsA); genes regulating hydrophobic cover formation (aerial mycelium) (bldB, bldC, bldM, bldN, sapA, chpC, chpD, chpE, chpH, ramA, ramC, ramS); and even some genes regulating early stages of sporulation (wblA, whiG, whiH, whiJ). The two most important differences between transcriptomes from liquid and solid cultures were: first, genes related to secondary metabolite biosynthesis (CDA, CPK, coelichelin, desferrioxamine clusters) were highly up-regulated in liquid but not in solid cultures; and second, genes involved in the final stages of hydrophobic cover/spore maturation (chpF, rdlA, whiE, sfr) were up-regulated in solid but not in liquid cultures. New information was also provided for several non-characterized genes differentially expressed in liquid and solid cultures which might be regulating, at least in part, the metabolic and developmental differences observed between liquid and solid cultures.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0086296PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3897704PMC
September 2014

Mycelium differentiation and development of Streptomyces coelicolor in lab-scale bioreactors: programmed cell death, differentiation, and lysis are closely linked to undecylprodigiosin and actinorhodin production.

Bioresour Technol 2014 Jan 30;151:191-8. Epub 2013 Oct 30.

Área de Microbiología, Departamento de Biología Funcional e IUOPA, Facultad de Medicina, Universidad de Oviedo, 33006 Oviedo, Spain.

Streptomycetes are mycelium-forming bacteria that produce two thirds of clinically relevant secondary metabolites. Secondary metabolite production is activated at specific developmental stages of Streptomyces life cycle. Despite this, Streptomyces differentiation in industrial bioreactors tends to be underestimated and the most important parameters managed are only indirectly related to differentiation: modifications to the culture media, optimization of productive strains by random or directed mutagenesis, analysis of biophysical parameters, etc. In this work the relationship between differentiation and antibiotic production in lab-scale bioreactors was defined. Streptomyces coelicolor was used as a model strain. Morphological differentiation was comparable to that occurring during pre-sporulation stages in solid cultures: an initial compartmentalized mycelium suffers a programmed cell death, and remaining viable segments then differentiate to a second multinucleated antibiotic-producing mycelium. Differentiation was demonstrated to be one of the keys to interpreting biophysical fermentation parameters and to rationalizing the optimization of secondary metabolite production in bioreactors.
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http://dx.doi.org/10.1016/j.biortech.2013.10.068DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3858829PMC
January 2014

Transcriptomic analysis of Streptomyces coelicolor differentiation in solid sporulating cultures: first compartmentalized and second multinucleated mycelia have different and distinctive transcriptomes.

PLoS One 2013 28;8(3):e60665. Epub 2013 Mar 28.

Área de Microbiología, Departamento de Biología Funcional and Instituto Universitario de Biotecnología de Asturias, Universidad de Oviedo, Oviedo, Spain.

Streptomycetes are very important industrial bacteria, which produce two thirds of all clinically relevant secondary metabolites. They have a complex developmental-cycle in which an early compartmentalized mycelium (MI) differentiates to a multinucleated mycelium (MII) that grows inside the culture medium (substrate mycelium) until it starts to growth into the air (aerial mycelium) and ends up forming spores. Streptomyces developmental studies have focused mainly on the later stages of MII differentiation (aerial mycelium and sporulation), with regulation of pre-sporulation stages (MI/MII transition) essentially unknown. This work represents the first study of the Streptomyces MI transcriptome, analyzing how it differs from the MII transcriptome. We have used a very conservative experimental approach to fractionate MI from MII and quantify gene expressions. The expression of well characterized key developmental/metabolic genes involved in bioactive compound production (actinorhodin, undecylprodigiosin, calcium-dependent antibiotic, cpk, geosmin) or hydrophobic cover formation-sporulation (bld, whi, wbl, rdl, chp, ram) was correlated with MII differentiation. Additionally, 122 genes conserved in the Streptomyces genus, whose biological function had not been previously characterized, were found to be differentially expressed (more than 4-fold) in MI or MII. These genes encoded for putative regulatory proteins (transcriptional regulators, kinases), as well as hypothetical proteins. Knowledge about differences between the MI (vegetative) and MII (reproductive) transcriptomes represents a huge advance in Streptomyces biology that will make future experiments possible aimed at characterizing the biochemical pathways controlling pre-sporulation developmental stages and activation of secondary metabolism in Streptomyces.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0060665PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3610822PMC
September 2013

Pre-sporulation stages of Streptomyces differentiation: state-of-the-art and future perspectives.

FEMS Microbiol Lett 2013 May 12;342(2):79-88. Epub 2013 Apr 12.

Área de Microbiología, Departamento de Biología Funcional, and IUBA, Facultad de Medicina, Universidad de Oviedo, Oviedo, Spain.

Streptomycetes comprise very important industrial bacteria, producing two-thirds of all clinically relevant secondary metabolites. They are mycelial microorganisms with complex developmental cycles that include programmed cell death (PCD) and sporulation. Industrial fermentations are usually performed in liquid cultures (large bioreactors), conditions in which Streptomyces strains generally do not sporulate, and it was traditionally assumed that there was no differentiation. In this work, we review the current knowledge on Streptomyces pre-sporulation stages of Streptomyces differentiation.
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http://dx.doi.org/10.1111/1574-6968.12128DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3654496PMC
May 2013

New Insights in Fermentations.

Ferment Technol 2012 Apr;1(2)

Area of Microbiology, Department of Functional Biology and IUBA, Faculty of Medicine, University of Oviedo, 33006 Oviedo, Spain.

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http://dx.doi.org/10.4172/2167-7972.1000e105DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3797508PMC
April 2012

New method for monitoring programmed cell death and differentiation in submerged Streptomyces cultures.

Appl Environ Microbiol 2010 May 26;76(10):3401-4. Epub 2010 Mar 26.

Area de Microbiologia, Departamento de Biologia Funcional de IUBA, Facultad de Medicina, Universidad de Oviedo, 33006 Oviedo, Spain.

Vital stains were used in combination with fluorimetry for the elaboration of a new method to quantify Streptomyces programmed cell death, one of the key events in Streptomyces differentiation. The experimental approach described opens the possibility of designing online protocols for automatic monitoring of industrial fermentations.
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http://dx.doi.org/10.1128/AEM.00120-10DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2869136PMC
May 2010

Mycelium differentiation and antibiotic production in submerged cultures of Streptomyces coelicolor.

Appl Environ Microbiol 2008 Jun 25;74(12):3877-86. Epub 2008 Apr 25.

Area de Microbiologia, Departamento de Biologia Funcional and IUBA, Facultad de Medicina, Universidad de Oviedo, 33006 Oviedo, Spain.

Despite the fact that most industrial processes for secondary metabolite production are performed with submerged cultures, a reliable developmental model for Streptomyces under these culture conditions is lacking. With the exception of a few species which sporulate under these conditions, it is assumed that no morphological differentiation processes take place. In this work, we describe new developmental features of Streptomyces coelicolor A3(2) grown in liquid cultures and integrate them into a developmental model analogous to the one previously described for surface cultures. Spores germinate as a compartmentalized mycelium (first mycelium). These young compartmentalized hyphae start to form pellets which grow in a radial pattern. Death processes take place in the center of the pellets, followed by growth arrest. A new multinucleated mycelium with sporadic septa (second mycelium) develops inside the pellets and along the periphery, giving rise to a second growth phase. Undecylprodigiosin and actinorhodin antibiotics are produced by this second mycelium but not by the first one. Cell density dictates how the culture will behave in terms of differentiation processes and antibiotic production. When diluted inocula are used, the growth arrest phase, emergence of a second mycelium, and antibiotic production are delayed. Moreover, pellets are less abundant and have larger diameters than in dense cultures. This work is the first to report on the relationship between differentiation processes and secondary metabolite production in submerged Streptomyces cultures.
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http://dx.doi.org/10.1128/AEM.02715-07DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2446541PMC
June 2008