GlnR-Mediated Regulation of ectABCD Transcription Expands the Role of the GlnR Regulon to Osmotic Stress Management

Abstract

Ectoine and hydroxyectoine are excellent compatible solutes for bacteria to deal with environmental osmotic stress and temperature damages. The biosynthesis cluster of ectoine and hydroxyectoine is widespread among microorganisms, and its expression is activated by high salinity and temperature changes. So far, little is known about the mechanism of the regulation of the transcription of ect genes and only two MarR family regulators (EctR1 in methylobacteria and the EctR1-related regulator CosR in Vibrio cholerae) have been found to negatively regulate the expression of ect genes. Here, we characterize GlnR, the global regulator for nitrogen metabolism in actinomycetes, as a negative regulator for the transcription of ectoine/hydroxyectoine biosynthetic genes (ect operon) in Streptomyces coelicolor. The physiological role of this transcriptional repression by GlnR is proposed to protect the intracellular glutamate pool, which acts as a key nitrogen donor for both the nitrogen metabolism and the ectoine/ hydroxyectoine biosynthesis. IMPORTANCEHigh salinity is deleterious, and cells must evolve sophisticated mechanisms to cope with this osmotic stress. Although production of ectoine and hydroxyectoine is one of the most frequently adopted strategies, the in-depth mechanism of regulation of their biosynthesis is less understood. So far, only two MarR family negative regulators, EctR1 and CosR, have been identified in methylobacteria and Vibrio, respectively. Here, our work demonstrates that GlnR, the global regulator for nitrogen metabolism, is a negative transcriptional regulator for ect genes in Streptomyces coelicolor. Moreover, a close relationship is found between nitrogen metabolism and osmotic resistance, and GlnR-mediated regulation of ect transcription is proposed to protect the intracellular glutamate pool. Meanwhile, the work reveals the multiple roles of GlnR in bacterial physiology. Ectoine and its hydroxyl derivative hydroxyectoine are the most widely used compatible solutes in bacteria to cope with environmental high osmotic stress (1-4). Because these molecules are small soluble organic compounds, they can prevent water loss and promote water reentry into the cells (3, 5). In addition, these compounds can also help stabilize proteins and protect bacteria from heat (6, 7) and cold (8, 9) damage. Also, with their excellent behavior in protein stabilization, ectoine and hydroxyectoine have been adopted in many industrial applications, e.g., in scientific research, cosmetics, and medical applications (3, 10-13).So far, the biosynthetic pathway of ectoine and hydroxyectoine has been thoroughly studied, and a total of four genes, ectABCD, are found to be involved in the processes (1,6,14,15). Specifically, L-aspartate-␤-semialdehyde is catalyzed into ectoine by a threestep enzymatic reaction, employing L-2,4-diaminobutyric acid transaminase (EctB), N-␥-acetyltransferase (EctA), and ectoine synthase (EctC). In some bacteria that contain ectD, encoding the ectoine hydroxylase, ectoine is hydroxylate...

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

“…The act and red genes were repressed, and genes for the TW95a pigment, and coelymicin were overexpressed. The genes for ectoine biosynthesis, controlled by GlnR (Shao et al, 2015 ), were overexpressed at all time points.…”

Insight into the Molecular Mechanism of the Transcriptional Regulation of amtB Operon in Streptomyces coelicolor

“…Recombinant S. coelicolor GlnR was expressed in E. coli BL21(DE3) and purification process was the same as previously described ( Shao et al, 2015 ). To prepare 6-carboxyfluorescein (FAM)-labeled probes for electrophoretic mobility shift assay (EMSA), primer FAM-M13F-47 was annealed with synthesized amtB oligos, followed by strand extension with Taq DNA polymerase (Tolo Biotech).…”

Post-translational Serine/Threonine Phosphorylation and Lysine Acetylation: A Novel Regulatory Aspect of the Global Nitrogen Response Regulator GlnR in S. coelicolor M145

“…Over 50 proteins associated with amino acid biosynthesis and carbon metabolism were shown to be differentially expressed between S. coelicolor M145 and the glnR mutant (Tiffert et al, 2011 ). GlnR-mediated control of carbohydrate transport (Liao et al, 2015 ) and regulation of ectoin (Shao et al, 2015 ) as well as validomycin A production (Qu et al, 2015 ) extended the GlnR role beyond the regulation of the N -assimilation. Regulation of the N -metabolism in S. coelicolor is very complex and depending on the conditions, involves additional control by other regulators such as: GlnRII (Fink et al, 2002 ; Reuther and Wohlleben, 2007 ), PhoP (Rodríguez-García et al, 2009 ; Martín et al, 2011 ; Sola-Landa et al, 2013 ), Crp (Gao et al, 2012 ), and AfsQ1 (Wang R. et al, 2013 ).…”