Responses of Mn2+speciation inDeinococcus radioduransandEscherichia colito γ-radiation by advanced paramagnetic resonance methods

Sharma, Ajay; Gaidamakova, Elena K.; Matrosova, Vera Y.; Bennett, Brian; Daly, Michael J.; Hoffman, Brian M.

doi:10.1073/pnas.1303376110

Cited by 70 publications

(102 citation statements)

References 36 publications

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“…19 By contrast, a separate study using similar EPR techniques concluded that such cells had little or no Mn(II) bound to proteins. 20 This study also demonstrated that in exponential cells Mn(II) speciation did not change after exposure to 10 kGy. 20 Without doubt as the D. radiodurans approaches stationary phase, Mn(II) speciation strongly shifts towards MnSOD and a second species characterized by a Mn(II) binding-site having two histidines, which we will refer to as Mn:(N-His) 2 .…”

Section: Introductionmentioning

confidence: 75%

The effect of gamma-ray irradiation on the Mn(ii) speciation in Deinococcus radiodurans and the potential role of Mn(ii)-orthophosphates

Bruch

Groot

et al. 2015

Metallomics

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D. radiodurans accumulates large quantities of Mn(II), which is believed to form low molecular weight complexes with phosphate and metabolites that protect D. radiodurans from radiation damage. The concentration of Mn(II) species in D. radiodurans during the exponential and stationary phase was determined using high-field EPR and biochemical techniques. In the exponential growth phase cells a large fraction of the manganese was in the form of Mn(II)-orthophosphate complexes. By contrast, the intracellular concentration of these compounds in stationary phase cells was less than 16 μM, while that of Mn superoxide dismutase was 320 μM and that of another, yet unidentified, Mn(II) protein was 250 μM. Stationary cells were found to be equally resistant to irradiation as the exponential cells in spite of having significant lower Mn(II)-orthophosphate concentrations. Gamma irradiation induced no changes in the Mn(II) speciation. During stationary growth phase D. radiodurans favours the production of the two Mn-proteins over low molecular weight complexes suggesting that the latter were not essential for radio-resistance at this stage of growth.

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Section: Introductionmentioning

confidence: 75%

The effect of gamma-ray irradiation on the Mn(ii) speciation in Deinococcus radiodurans and the potential role of Mn(ii)-orthophosphates

Bruch

Groot

et al. 2015

Metallomics

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“…Importantly, manganese accumulation was also observed during stress response in other bacterial phyla as well as in eukaryotic organisms (44). Although the data on intracellular distribution of Mn 2+ ions in Dra are controversial (26,(45)(46)(47), several reports suggest that a significant fraction of Mn 2+ ions is bound to proteins (46,48). Accordingly, Mn 2+ ions were shown to activate several stressinduced enzymes in Dra, including DNA polymerases (49,50), Mn 2+ -dependent superoxide dismutase (45,48), and the PprI protease responsible for activation of dozens of stress-related genes through the cleavage of DdrO repressor (51).…”

Section: Discussionmentioning

confidence: 99%

Regulation of transcriptional pausing through the secondary channel of RNA polymerase

Esyunina

Agapov

Kulbachinskiy

2016

Proc. Natl. Acad. Sci. U.S.A.

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Transcriptional pausing has emerged as an essential mechanism of genetic regulation in both bacteria and eukaryotes, where it serves to coordinate transcription with other cellular processes and to activate or halt gene expression rapidly in response to external stimuli. Deinococcus radiodurans, a highly radioresistant and stressresistant bacterium, encodes three members of the Gre family of transcription factors: GreA and two Gre factor homologs, Gfh1 and Gfh2. Whereas GreA is a universal bacterial factor that stimulates RNA cleavage by RNA polymerase (RNAP), the functions of lineage-specific Gfh proteins remain unknown. Here, we demonstrate that these proteins, which bind within the RNAP secondary channel, strongly enhance site-specific transcriptional pausing and intrinsic termination. Uniquely, the pause-stimulatory activity of Gfh proteins depends on the nature of divalent ions (Mg 2+ or Mn 2+ ) present in the reaction and is also modulated by the nascent RNA structure and the trigger loop in the RNAP active site. Our data reveal remarkable plasticity of the RNAP active site in response to various regulatory stimuli and highlight functional diversity of transcription factors that bind inside the secondary channel of RNAP.RNA polymerase | transcriptional pausing | Gfh factors | Deinococcus radiodurans | stress response C ellular RNA polymerases (RNAPs) are complex molecular machines whose activity during transcription is regulated by DNA-and RNA-encoded signals, protein factors, small molecules, and inhibitors. The catalytic cycle of RNAP can be interrupted by pauses of various natures that play important roles in genetic regulation in all organisms, from the classic systems of transcription attenuation and their variations in bacteria (1) to recently discovered widespread promoter-proximal pausing in eukaryotes (2). The pausing serves to activate or repress transcription rapidly at specific genomic sites in response to regulatory stimuli and to coordinate RNA synthesis with other genetic processes (e.g., DNA replication and repair, RNA translation in bacteria) (1,(3)(4)(5)(6)(7)(8).Recent structural and biochemical studies revealed distinct RNAP conformations corresponding to different functional states of the transcription elongation complex (TEC) during nucleotide addition, RNA proofreading, and pausing (9-11). The control of structural transitions between these states likely underlies the function of multiple regulatory factors acting on RNAP. However, the roles of individual RNAP conformations in transcription and the mechanisms of their switching by transcription factors remain only partially understood. During transcription, RNAP holds the DNA template and the RNA transcript within its main channel, whose opening is controlled by the mobile clamp/shelf module and the flap domain of RNAP (9-11). Nucleotide addition and TEC translocation depend on alternating cycles of the folding of the trigger loop (TL) and kinking of the bridge helix (BH) in the RNAP active site (9-11) (Fig. 1A). Analysis of the stru...

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“…The labile manganese pool is likely buffered as an LMW pool bound to phosphate, nucleotides, peptides, and organic acids (24,25), including the abundant metabolite fructose 1,6-diphosphate (26). The labile iron pool is thought to be largely chelated by LMW thiols such as glutathione, with a lesser contribution from citrate and other organic acids (27).…”

Section: Metal Ion Speciation In the Cellmentioning

confidence: 99%

Specificity of Metal Sensing: Iron and Manganese Homeostasis in Bacillus subtilis

Helmann

2014

Journal of Biological Chemistry

129

127

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Metalloregulatory proteins allow cells to sense metal ions and appropriately adjust the expression of metal uptake, storage, and efflux pathways. Bacillus subtilis provides a model for the coordinate regulation of iron and manganese homeostasis that involves three key regulators: Fur senses iron sufficiency, MntR senses manganese sufficiency, and PerR senses the intracellular Fe/Mn ratio. Here, I review the structural and physiological bases of selective metal perception, the effects of non-cognate metals, and mechanisms that may serve to coordinate iron and manganese homeostasis. Manganese and Iron: Chemically Similar Elements with Distinct Roles in Cell PhysiologyManganese and iron are both predominantly present as divalent cations in the reducing environment of the cytoplasm and are maintained at overall concentrations (averaged over the cell) that can approach 0.5 mM (1-3). Much of this manganese and iron is bound by enzymes that acquire metal from a kinetically accessible (labile) pool buffered in the low M range (4 -6). These two metals are not only important from the perspective of individual microbial cells, but have a global impact due to their central roles in photosynthesis, nitrogen fixation, and other key steps in elemental cycling.Nearly all cellular life requires significant amounts of iron. The sole documented exceptions are the lactobacilli and the spirochete Borrelia burgdorferi, which have little if any requirement for iron (3). Coincident with this disregard for iron, these same organisms have high requirements for manganese and have been described as manganese-centric (7). Conversely, other organisms have little demonstrable requirement for manganese, although they may use manganese when available. The best characterized is Escherichia coli, which has a largely ironcentric metabolism, but conditionally imports manganese in response to oxidative stress (8).Here, I review the key factors regulating iron and Mn homeostasis in the model organism Bacillus subtilis. B. subtilis encodes three members of the Fur 2 family (Fur, PerR, and Zur) and one member of the DtxR/MntR family (MntR) that regulate the import of nutrient metal ions (9 -11). Structural and biochemical studies have provided insights into the mechanisms of selective activation of these repressors by their cognate metal ions (12-14). B. subtilis requires both iron and manganese for growth, a property shared with many pathogens for which the ability to obtain these metals from the host is a critical determinant of virulence (7,(15)(16)(17). Metalloregulators as a Window into Cellular PhysiologyMetalloregulators function as the arbiters of metal ion sufficiency and excess (6,12,18). For those that sense sufficiency, the affinity (measured as K d ) for their cognate metal(s) defines the level of metal judged to be sufficient. When free metal concentrations rise above this level, the regulator transitions from its inactive (apo-) form to its activated (holo-) form and represses expression of metal import. In B. subtilis, Fur serves as ...

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Responses of Mn²⁺speciation inDeinococcus radioduransandEscherichia colito γ-radiation by advanced paramagnetic resonance methods

Cited by 70 publications

References 36 publications

The effect of gamma-ray irradiation on the Mn(ii) speciation in Deinococcus radiodurans and the potential role of Mn(ii)-orthophosphates

The effect of gamma-ray irradiation on the Mn(ii) speciation in Deinococcus radiodurans and the potential role of Mn(ii)-orthophosphates

Regulation of transcriptional pausing through the secondary channel of RNA polymerase

Specificity of Metal Sensing: Iron and Manganese Homeostasis in Bacillus subtilis

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