Transcriptional regulation of the Pseudomonas aeruginosa iron-sulfur cluster assembly pathway by binding of IscR to multiple sites

Abstract

Iron-sulfur ([Fe-S]) cluster proteins have essential functions in many biological processes. [Fe-S] homeostasis is crucial for bacterial survival under a wide range of environmental conditions. IscR is a global transcriptional regulator in Pseudomonas aeruginosa ; it has been shown to regulate genes involved in [Fe-S] cluster biosynthesis, iron homeostasis, resistance to oxidants, and pathogenicity. Many aspects of the IscR transcriptional regulatory mechanism differ from those of other well-studied systems. This study demonstrates the mechanisms of IscR Type-1 binding to its target sites that mediate the repression of gene expression at the isc operon, nfuA , and tpx . The analysis of IscR binding to multiple binding sites in the promoter region of the isc operon reveals that IscR first binds to the high-affinity site B followed by binding to the low-affinity site A. The results of in vitro IscR binding assays and in vivo analysis of IscR-mediated repression of gene expression support the role of site B as the primary site, while site A has only a minor role in the efficiency of IscR repression of gene expression. Ligation of an [Fe-S] cluster to IscR is required for the binding of IscR to target sites and in vivo repression and stress-induced gene expression. Analysis of Type-1 sites in many bacteria, including P . aeruginosa , indicates that the first and the last three AT-rich bases were among the most highly conserved bases within all analyzed Type-1 sites. Herein, we first propose the putative sequence of P . aeruginosa IscR Type-1 binding motif as . This can benefit further studies in the identification of novel genes under the IscR regulon and the regulatory mechanism model of P . aeruginosa IscR as it contributes to the roles of an [Fe-S] cluster in several biologically important cellular activities.

Genomic Characterization of Carbapenem-Non-susceptible Pseudomonas aeruginosa Clinical Isolates From Saudi Arabia Revealed a Global Dissemination of GES-5-Producing ST235 and VIM-2-Producing ST233 Sub-Lineages

“…GES-positive genomes were separated from the remaining ST235 genomes by at least 29 SNPs, of which, 14 were non-synonymous in genes classified as transcriptional regulators, metabolic genes and hypothetical proteins whereas four were located in the promoter or potential regulatory regions of genes encoding the porin OprO, global transcriptional regulator IscR or hypothetical proteins ( Supplementary Table 2 ). Of these, only IscR has been shown to regulate genes involved in iron homeostasis, resistance to oxidants and pathogenicity ( Romsang et al, 2014 ; Saninjuk et al, 2019 ). Comparison of the gene contents of ST235 genomes identified 4,373 core and 1,086 soft-core genes that were shared among 98.94–100% and 94.87–98.94% of genomes studied, respectively.…”

Oxidative Stress Response in Pseudomonas aeruginosa

“…IscR (iron–sulfur cluster) is a transcriptional regulator that belongs to the Rrf2 family and regulates the expression of genes involved in iron–sulfur ([Fe–S]) cluster biosynthesis. [Fe–S] cluster proteins play essential roles in several cellular processes, including respiration, gene regulation, DNA repair, electron storage, and central metabolism [ 206 , 207 , 208 ]. In P. aeruginosa only the isc system encoded by the iscRSUA-hscBA-fdx2-iscX operon has been identified [ 110 ].…”

“…[Fe-S] cluster proteins play essential roles in several cellular processes, including respiration, gene regulation, DNA repair, electron storage, and central metabolism [206][207][208]. In P. aeruginosa only the isc system encoded by the iscRSUA-hscBA-fdx2-iscX operon has been identified [110].…”

Regulatory and Sensing Iron-Sulfur Clusters: New Insights and Unanswered Questions

“…Mammalian, bacterial, and yeast cells utilize Fe-S cluster containing iron regulatory proteins for intracellular iron sensing. In the broadest sense, iron regulatory proteins, such as IRP1 (aka ACO1) in humans [1,7,10,102,[134][135][136][137], IscR/RirA in bacteria [8,27,[138][139][140][141][142], and Aft1 in yeast [143][144][145][146] sense iron levels via the presence or absence of Fe-S clusters. In iron replete conditions, ligation of Fe-S cluster cofactors is favored.…”