title: WhiB7, an Fe-S-dependent transcription factor that activates species-specific repertoires of drug resistance determinants in actinobacteria.
abstract: WhiB-like (Wbl) proteins are well known for their diverse roles in actinobacterial morphogenesis, cell division, virulence, primary and secondary metabolism, and intrinsic antibiotic resistance. Gene disruption experiments showed that three different Actinobacteria (Mycobacterium smegmatis, Streptomyces lividans, and Rhodococcus jostii) each exhibited a different whiB7-dependent resistance profile. Heterologous expression of whiB7 genes showed these resistance profiles reflected the host's repertoire of endogenous whiB7-dependent genes. Transcriptional activation of two resistance genes in the whiB7 regulon, tap (a multidrug transporter) and erm(37) (a ribosomal methyltransferase), required interaction of WhiB7 with their promoters. Furthermore, heterologous expression of tap genes isolated from Mycobacterium species demonstrated that divergencies in drug specificity of homologous structural proteins contribute to the variation of WhiB7-dependent drug resistance. WhiB7 has a specific tryptophan/glycine-rich region and four conserved cysteine residues; it also has a peptide sequence (AT-hook) at its C terminus that binds AT-rich DNA sequence motifs upstream of the promoters it activates. Targeted mutagenesis showed that these motifs were required to provide antibiotic resistance in vivo. Anaerobically purified WhiB7 from S. lividans was dimeric and contained 2.1 ± 0.3 and 2.2 ± 0.3 mol of iron and sulfur, respectively, per protomer (consistent with the presence of a 2Fe-2S cluster). However, the properties of the dimer's absorption spectrum were most consistent with the presence of an oxygen-labile 4Fe-4S cluster, suggesting 50% occupancy. These data provide the first insights into WhiB7 iron-sulfur clusters as they exist in vivo, a major unresolved issue in studies of Wbl proteins.
authors.affiliation: From the Department of Microbiology and Immunology, Centre for Tuberculosis Research, Life Sciences Centre, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada.
authors.forename: Santiago
authors.initials: S
authors.lastname: Ramón-García
authors.forename: Carol
authors.initials: C
authors.lastname: Ng
authors.forename: Pernille R
authors.initials: PR
authors.lastname: Jensen
authors.forename: Manisha
authors.initials: M
authors.lastname: Dosanjh
authors.forename: Jan
authors.initials: J
authors.lastname: Burian
authors.forename: Rowan P
authors.initials: RP
authors.lastname: Morris
authors.forename: Marc
authors.initials: M
authors.lastname: Folcher
authors.forename: Lindsay D
authors.initials: LD
authors.lastname: Eltis
authors.forename: Stephan
authors.initials: S
authors.lastname: Grzesiek
authors.forename: Liem
authors.initials: L
authors.lastname: Nguyen
authors.forename: Charles J
authors.initials: CJ
authors.lastname: Thompson
mesh_terms: D039903:Actinobacteria; D001426:Bacterial Proteins / Q000235:genetics*; D024881:Drug Resistance, Bacterial / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D054884:Host-Pathogen Interactions / Q000235:genetics; D007506:Iron-Sulfur Proteins / Q000235:genetics* / Q000378:metabolism; D008780:Methyltransferases / Q000378:metabolism; D016296:Mutagenesis; D020102:Mycobacterium smegmatis / Q000235:genetics / Q000378:metabolism*; D012240:Rhodococcus / Q000235:genetics / Q000378:metabolism*; D013045:Species Specificity; D048170:Streptomyces lividans / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D015533:Transcriptional Activation / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D007506:Iron-Sulfur Proteins; D014157:Transcription Factors; D008780:Methyltransferases
keywords: AT-hook; Actinobacteria; Antibiotic Resistance; Gene Regulation; Iron-Sulfur Cluster; Iron-Sulfur Protein; Regulation; Transcription