title: Tryptophan promotes morphological and physiological differentiation in Streptomyces coelicolor.
abstract: The molecular mechanisms regulating tryptophan biosynthesis in actinomycetes are poorly understood; similarly, the possible roles of tryptophan in the differentiation program of microorganism life-cycle are still underexplored. To unveil the possible regulatory effect of this amino acid on gene expression, an integrated study based on quantitative teverse transcription-PCR (qRT-PCR) and proteomic approaches was performed on the actinomycete model Streptomyces coelicolor. Comparative analyses on the microorganism growth in a minimal medium with or without tryptophan supplementation showed that biosynthetic trp gene expression in S. coelicolor is not subjected to a negative regulation by the presence of the end product. Conversely, tryptophan specifically induces the transcription of trp genes present in the biosynthetic gene cluster of the calcium-dependent antibiotic (CDA), a lipopeptide containing D- and L-tryptophan residues. In addition, tryptophan stimulates the transcription of the CDA gene cluster regulator cdaR and, coherently, CDA production. Surprisingly, tryptophan also promotes the production of actinorhodin, another antibiotic that does not contain this amino acid in its structure. Combined 2D-DIGE and nano liquid chromatography electrospray linear ion trap tandem mass spectrometry (LC-ESI-LIT-MS/MS) analyses revealed that tryptophan exerts a growth-stage-dependent global effect on S. coelicolor proteome, stimulating anabolic pathways and promoting the accumulation of key factors associated with morphological and physiological differentiation at the late growth stages. Phenotypic observations by scanning electron microscopy and spore production assays demonstrated an increased sporulation in the presence of tryptophan. Transcriptional analysis of catabolic genes kynA and kynB suggested that the actinomycete also uses tryptophan as a carbon and nitrogen source. In conclusion, this study originally provides the molecular basis underlying the stimulatory effect of tryptophan on the production of antibiotics and morphological development program of this actinomycete.
authors.affiliation: Laboratory of Genetics, School of Biosciences and Veterinary Medicine, University of Camerino, 62032, Camerino, Italy. emilia.palazzotto@unicam.it.|Laboratory of Molecular Microbiology and Biotechnology, STEBICEF Department, University of Palermo, 90128, Palermo, Italy. emilia.palazzotto@unicam.it.
authors.forename: Emilia
authors.initials: E
authors.lastname: Palazzotto
authors.affiliation: Proteomic and Mass Spectrometry Laboratory, ISPAAM, National Research Council, 80147, Naples, Italy.
authors.forename: Giovanni
authors.initials: G
authors.lastname: Renzone
authors.affiliation: Laboratory of Molecular Microbiology and Biotechnology, STEBICEF Department, University of Palermo, 90128, Palermo, Italy.
authors.forename: Pietro
authors.initials: P
authors.lastname: Fontana
authors.affiliation: Dipartimento di Ingegneria Civile, Ambientale, Aerospaziale, dei Materiali, Università di Palermo, 90128, Palermo, Italy.
authors.forename: Luigi
authors.initials: L
authors.lastname: Botta
authors.affiliation: Proteomic and Mass Spectrometry Laboratory, ISPAAM, National Research Council, 80147, Naples, Italy.
authors.forename: Andrea
authors.initials: A
authors.lastname: Scaloni
authors.affiliation: Laboratory of Molecular Microbiology and Biotechnology, STEBICEF Department, University of Palermo, 90128, Palermo, Italy.
authors.forename: Anna Maria
authors.initials: AM
authors.lastname: Puglia
authors.affiliation: Laboratory of Molecular Microbiology and Biotechnology, STEBICEF Department, University of Palermo, 90128, Palermo, Italy.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Gallo
mesh_terms: D002244:Carbon / Q000378:metabolism; D002853:Chromatography, Liquid; D003470:Culture Media / Q000737:chemistry; D015180:Electrophoresis, Gel, Two-Dimensional; D004734:Energy Metabolism; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects*; D008855:Microscopy, Electron, Scanning; D009584:Nitrogen / Q000378:metabolism; D020543:Proteome / Q000032:analysis; D021241:Spectrometry, Mass, Electrospray Ionization; D013171:Spores, Bacterial / Q000254:growth & development / Q000648:ultrastructure; D048372:Streptomyces coelicolor / Q000166:cytology* / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D014364:Tryptophan / Q000378:metabolism*
chemical_list: D003470:Culture Media; D020543:Proteome; D002244:Carbon; D014364:Tryptophan; D009584:Nitrogen
keywords: 2D-DIGE; Actinorhodin; CDA; Differentiation; S. coelicolor; Tryptophan