Sequential binding of FurA from Anabaena sp. PCC 7120 to iron boxes: Exploring regulation at the nanoscale

Pallarés, María Carmen; Marcuello, Carlos; Botello-Morte, Laura; González, Andrés; Fillat, Marı́a F.; Lostao, Anabel

doi:10.1016/j.bbapap.2014.01.005

Cited by 14 publications

(16 citation statements)

References 63 publications

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“…Biomolecules were then immobilized on freshly cleaved muscovite mica sheets (Electron Microscopy Sciences). Mica pieces were pretreated with 200 mM MgCl 2 for 5 minutes and rinsed with binding buffer prior to biomolecule binding as optimised previously to slightly immobilise DNA strands on such negative surfaces [30]. The concentration of the DNA incubated on the substrates was suitable to get isolated complexes that could be analysed individually.…”

Section: Atomic Force Microscopy (Afm)mentioning

confidence: 99%

Molecular basis for the integration of environmental signals by FurB from Anabaena sp. PCC 7120

Sein‐Echaluce

Pallarés

Lostao

et al. 2018

Biochemical Journal

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FUR (Ferric uptake regulator) proteins are among the most important families of transcriptional regulators in prokaryotes, often behaving as global regulators. In the cyanobacterium PCC 7120, FurB (Zur, Zinc uptake regulator) controls zinc and redox homeostasis through the repression of target genes in a zinc-dependent manner., non-specific binding of FurB to DNA elicits protection against oxidative damage and avoids cleavage by deoxyribonuclease I. The present study provides, for the first time, evidence of the influence of redox environment in the interaction of FurB with regulatory zinc and its consequences in FurB-DNA-binding affinity. Calorimetry studies showed that, in addition to one structural Zn(II), FurB is able to bind two additional Zn(II) per monomer and demonstrated the implication of cysteine C93 in regulatory Zn(II) coordination. The interaction of FurB with the second regulatory zinc occurred only under reducing conditions. While non-specific FurB-DNA interaction is Zn(II)-independent, the optimal binding of FurB to target promoters required loading of two regulatory zinc ions. Those results combined with site-directed mutagenesis and gel-shift assays evidenced that the redox state of cysteine C93 conditions the binding of the second regulatory Zn(II) and, in turn, modulates the affinity for a specific DNA target. Furthermore, differential spectroscopy studies showed that cysteine C93 could also be involved in heme coordination by FurB, either as a direct ligand or being located near the binding site. The results indicate that besides controlling zinc homeostasis, FurB could work as a redox-sensing protein probably modifying its zinc and DNA-binding abilities depending upon environmental conditions.

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Section: Atomic Force Microscopy (Afm)mentioning

confidence: 99%

Molecular basis for the integration of environmental signals by FurB from Anabaena sp. PCC 7120

Sein‐Echaluce

Pallarés

Lostao

et al. 2018

Biochemical Journal

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“…Because of their ability to oligomerize in a metal-dependent fashion1213 and to bind promoters at multiple sites141516, they represent ideal candidates for the investigation of metal-dependent DNAe. In addition, topological modifications of the DNA induced by Fur binding, such as bending and wrapping, have been reported in vitro using various footprinting and microscopy techniques, including atomic force microscopy (AFM)1718.…”

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confidence: 99%

Metal-responsive promoter DNA compaction by the ferric uptake regulator

et al. 2016

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Short-range DNA looping has been proposed to affect promoter activity in many bacterial species and operator configurations, but only few examples have been experimentally investigated in molecular detail. Here we present evidence for a metal-responsive DNA condensation mechanism controlled by the Helicobacter pylori ferric uptake regulator (Fur), an orthologue of the widespread Fur family of prokaryotic metal-dependent regulators. H. pylori Fur represses the transcription of the essential arsRS acid acclimation operon through iron-responsive oligomerization and DNA compaction, encasing the arsR transcriptional start site in a repressive macromolecular complex. A second metal-dependent regulator NikR functions as nickel-dependent anti-repressor at this promoter, antagonizing the binding of Fur to the operator elements responsible for the DNA condensation. The results allow unifying H. pylori metal ion homeostasis and acid acclimation in a mechanistically coherent model, and demonstrate, for the first time, the existence of a selective metal-responsive DNA compaction mechanism controlling bacterial transcriptional regulation.

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“…This is supported by the observed up-regulation of the FurA repressor upon nitrogen deprivation, possibly through NtcA (Lopez-Gomollon et al 2007). FurA acts as a typical repressor requiring Fe 2+ as a co-repressor for binding effectively to the iron boxes (Pallares et al 2014). Under Fe 2+ -deficient conditions, the release of Fe 2+ from FurA results in the dissociation of the FurA from the iron boxes thereby derepressing the expression of the downstream genes.…”

Section: Discussionmentioning

confidence: 85%

“…FurA acts as a typical repressor requiring Fe 2+ as a co‐repressor for binding effectively to the iron boxes (Pallares et al. ). Under Fe 2+ ‐deficient conditions, the release of Fe 2+ from FurA results in the dissociation of the FurA from the iron boxes thereby derepressing the expression of the downstream genes.…”

Section: Discussionmentioning

confidence: 99%

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Differential regulation of ssb genes in the nitrogen‐fixing cyanobacterium, Anabaena sp. strain PCC7120¹

Kirti

Kumar

Rajaram

2017

Journal of Phycology

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Anabaena sp. PCC7120 possesses three genes coding for single-stranded DNA-binding (SSB) protein, of which ssb1 was a single gene, and ssb2 and ssb3 are the first genes of their corresponding operons. Regulation of the truncated ssb genes, ssb1 (alr0088) and ssb2 (alr7559), was unaffected by N-status of growth. They were negatively regulated by the SOS-response regulatory protein LexA, as indicated by the (i) binding of Anabaena LexA to the LexA box of regulatory regions of ssb1 and ssb2, and (ii) decreased expression of the downstream gfp reporter gene in Escherichia coli upon co-expression of LexA. However, the full-length ssb gene, ssb3 (all4779), was regulated by the availability of Fe and combined nitrogen, as indicated by (i) increase in the levels of SSB3 protein on Fe -depletion and decrease under Fe -excess conditions, and (ii) 1.5- to 1.6-fold decrease in activity under nitrogen-fixing conditions compared to nitrogen-supplemented conditions. The requirement of Fe as a co-factor for repression by FurA and the increase in levels of FurA under nitrogen-deficient conditions in Anabaena (Lopez-Gomollon et al. 2007) indicated a possible regulation of ssb3 by FurA. This was substantiated by (i) the binding of FurA to the regulatory region of ssb3, (ii) repression of the expression of the downstream gfp reporter gene in E. coli upon co-expression of FurA, and (iii) negative regulation of ssb3 promoter activity by the upstream AT-rich region in Anabaena. This is the first report on possible role of FurA, an important protein for iron homeostasis, in DNA repair of cyanobacteria.

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Sequential binding of FurA from Anabaena sp. PCC 7120 to iron boxes: Exploring regulation at the nanoscale

Cited by 14 publications

References 63 publications

Molecular basis for the integration of environmental signals by FurB from Anabaena sp. PCC 7120

Molecular basis for the integration of environmental signals by FurB from Anabaena sp. PCC 7120

Metal-responsive promoter DNA compaction by the ferric uptake regulator

Differential regulation of ssb genes in the nitrogen‐fixing cyanobacterium, Anabaena sp. strain PCC7120¹

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Sequential binding of FurA from Anabaena sp. PCC 7120 to iron boxes: Exploring regulation at the nanoscale

Cited by 14 publications

References 63 publications

Molecular basis for the integration of environmental signals by FurB from Anabaena sp. PCC 7120

Molecular basis for the integration of environmental signals by FurB from Anabaena sp. PCC 7120

Metal-responsive promoter DNA compaction by the ferric uptake regulator

Differential regulation of ssb genes in the nitrogen‐fixing cyanobacterium, Anabaena sp. strain PCC71201

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Differential regulation of ssb genes in the nitrogen‐fixing cyanobacterium, Anabaena sp. strain PCC7120¹