Identification of two‐component system AfsQ1/Q2 regulon and its cross‐regulation with GlnR in S treptomyces coelicolor

• Published in: Molecular microbiology Vol. 87; no. 1; pp. 30 - 48
• Main Authors: Wang, Rui, Mast, Yvonne, Wang, Jin, Zhang, Weiwen, Zhao, Guoping, Wohlleben, Wolfgang, Lu, Yinhua, Jiang, Weihong
• Format: Journal Article
• Language: English
• Published: 01.01.2013
• ISSN: 0950-382X; 1365-2958
• DOI: 10.1111/mmi.12080

The two‐component system AfsQ1/Q2 of S treptomyces coelicolor was identified in our previous work as a pleiotropic regulator for antibiotic biosynthesis and morphological differentiation under the condition of a minimal medium supplemented with 75 m M glutamate. In this work, we report the dissection of the mechanism underlying the function of AfsQ1/Q2 on antibiotic production and also the identification of the AfsQ1/Q2 regulon. The results showed that AfsQ1/Q2 stimulated antibiotic ACT , RED and CDA production directly through the pathway‐specific activator genes actII‐ORF4 , redZ and cdaR respectively. In addition, expression of sigQ that encodes a sigma factor and is divergently transcribed from afsQ1 was also subject to direct regulation by AfsQ1/Q2 . The precise AfsQ1 binding sites in the upstream regions of these target genes were determined by DN ase I footprinting assays coupled with site‐directed DNA mutagenesis. By computational prediction and functional analysis, at least 17 new AfsQ1 targets were identified, including pstS gene encoding a high‐affinity phosphate‐binding protein and two developmental genes whiD , bldM . For the AfsQ1/Q2 regulon, an AfsQ1 binding motif comprising the sequence GTnAC‐n 6 ‐GTnAC has been defined. Interestingly, we found from electrophoretic mobility shift assays and transcriptional analysis that AfsQ1/Q2 can also function as a repressor for nitrogen assimilation, and AfsQ1 can compete with GlnR for the promoter regions of glnA and nirB , suggesting the cross‐regulation between AfsQ1/Q2 and GlnR in nitrogen metabolism. These findings suggested that AfsQ1/Q2 is important not only for antibiotic biosynthesis but also in maintaining the metabolic homeostasis of nutrient utilization under the stress of high concentration of glutamate in S . coelicolor .