Expression, Functional Characterization and X-ray Analysis of HosA, A Member of MarR Family of Transcription Regulator from Uropathogenic Escherichia coli

Reddi, Ravikumar; Sawhney, Bhavik; Ghosh, Debasish Kumar; Addlagatta, A.; Ranjan, Akash

doi:10.1007/s10930-016-9670-1

Cited by 11 publications

(12 citation statements)

References 33 publications

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“…VanR relates to the PadR‐like transcriptional regulator family. Its binding site in C. glutamicum consists of inverted repeats (AACTAACTAA(N4)TTAGGTATTT) . The putative binding site of the PadR protein in S. fradiae is located 13 bpup stream from the start codon of the marR ‐family transcriptional regulator gene (KNE81702.1) and has a 70% identity with the known VanR protein‐binding site (Fig.…”

Section: Resultsmentioning

confidence: 99%

A functional study of the global transcriptional regulator PadR from a strain Streptomyces fradiae‐nitR+bld, resistant to nitrone‐oligomycin

et al. 2018

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We describe Streptomyces fradiae mechanisms of sensitivity to nitrone-oligomycin A, a derivative of oligomycin A. We obtained S. fradiae-nitR bld, a nitrone-oligomycin A resistant mutant with a «bald» phenotype. Comparative genomic analysis of the wild-type S. fradiae ATCC19609 and S. fradiae-nitR bld revealed a mutation in padR - a gene encoding a multifunction transcription regulator, which resulted in the amino acid replacement in a highly conserved DNA-binding domain. Bioinformatics genome analysis of S. fradiae ATCC19609 discovered a PadR binding site 13 bp upstream the start codon of the marR transcription factor gene. Induction of S. fradiaenitR bld and w.t. strains with nitrone-oligomycin A lead to a significant increase in expression level of the marR gene in the w.t. strain, but no change observed in mutant strain. We identified differences between DNA-protein interactions of the mutant and native PadR proteins with its putative binding site in S. fradiae ATCC19609. This allowed us to suggest that the padR gene, that harbored a single nucleotide mutation in the S. fradiaenitR bld strain, might be involved in the mechanism of resistance to nitrone-oligomycin A. We assume the participation of the transcriptional factorpadR in the formation of the bald phenotype.

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Section: Resultsmentioning

confidence: 99%

A functional study of the global transcriptional regulator PadR from a strain Streptomyces fradiae‐nitR+bld, resistant to nitrone‐oligomycin

et al. 2018

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“…MarR family regulators are widespread in prokaryotes, and members of MarR family of transcription regulators exhibit high structural similarity despite low sequence similarity [35]. The sequence dissimilarity might be required to respond to diverse signaling molecules and recognize unique DNA targets [35]. To verify the regulatory role, the ctcS gene was genetically interrupted firstly.…”

Section: Discussionmentioning

confidence: 99%

CtcS, a MarR family regulator, regulates chlortetracycline biosynthesis

et al. 2019

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BackgroundChlortetracycline (CTC) is one of the commercially important tetracyclines (TCs) family product and is mainly produced by Streptomyces. CTC is still in a great demand due to its broad-spectrum activity against pathogens. Engineering transcriptional control allows the cell to allocate its valuable resources towards protein production and provides an important method for the build-up of desired metabolites. Despite extensive efforts concerning transcriptional regulation for increasing the productivities of TCs, the regulatory mechanisms of the CTC biosynthesis remain poorly understood.ResultsIn this study, the possible regulatory function of CtcS, a potential member of MarR (multiple antibiotic resistance regulator) family of transcriptional regulators in S. aureofaciens F3, was demonstrated. Knockdown of ctcS altered the transcription of several biosynthesis-related genes and reduced the production of tetracycline (TC) and CTC, without obvious effect on morphological differentiation and cell growth. Especially, CtcS directly repressed the transcription of the adjacent divergent gene ctcR (which encodes a putative TC resistance efflux protein). A CtcS-binding site was identified within the promoter region of ctcR by DNase I footprinting and an inverted repeat (5′-CTTGTC-3′) composed of two 6-nt half sites in the protected region was found. Moreover, both CTC and TC could attenuate the binding activity of CtcS with target DNA.ConclusionctcS regulated the production of TC and CTC in S. aureofaciens F3 and the overexpression of it could be used as a simple approach for the construction of engineering strain with higher productivity. Meanwhile, CtcS was characterized as a TC- and CTC-responsive MarR family regulator. This study provides a previously unrecognized function of CtcS and will benefit the research on the regulatory machinery of the MarR family regulators.

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“…The HYPK, HYPK-UBA, HYPK-UBA D94A, E101A, HYPK-UBA ΔL113, ΔG118, HYPK-N60, HYPK-C69, HYPK-Δ 48 DYA 50 , LC3, Nedd8, Htt97Qexon1 recombinant proteins were produced from pET21b vectors by the T7 approach that was described by us in earlier studies ( 20, 42 ). In the pETDuet-1 vector system, HYPK/ HYPK-N60/ HYPK-C69/ HYPK-Δ 48 DYA 50 and LC3A/ LC3B/ GABARAP/ GABARAPL1/ GABARAPL2 were 6xHis tagged and the interacting proteins (LC3 and HYPK) were untagged.…”

Section: Methodsmentioning

confidence: 99%

HYPK scaffolds the Nedd8 and LC3 proteins to initiate formation of autophagosome around polyneddylated huntingtin exon1 aggregates

Ghosh

Ranjan

2019

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Selective autophagy of protein aggregates is necessary for maintaining the cellular proteostasis. Several regulatory proteins play critical roles in this process. Here, we report that the huntingtin interacting protein K (HYPK) modulates the autophagic degradation of poly-neddylated huntingtin exon1 aggregates. HYPK functions as a scaffolding protein that binds to the Nedd8 and LC3 proteins. The C-terminal ubiquitin-associated (UBA) domain of HYPK binds to the Nedd8, whereas an N-terminal tyrosine-type (Y-type) LC3 interacting region (LIR) of HYPK binds to the LC3. Several conserved amino acids in the UBA domain of HYPK are necessary to mediate the efficient binding of HYPK to Nedd8. The autophagy inducing properties of HYPK are manifested by the increased lipidation of LC3 protein, increased expression of beclin-1 and ATG-5 proteins, and generation of puncta-like granules of LC3 in the HYPK overexpressing cells. Association of the ‘H-granules’ of HYPK with the poly-neddylated huntingtin exon1 aggregates results in the formation of autophagosome around the huntingtin exon1 aggregates, thereby clearing the aggregates by aggrephagy. Poly-neddylation of huntingtin exon1 is required for its autophagic degradation by HYPK. Thus, overexpression of Nedd8 also increases the basal level of cellular autophagy, other than maintaining the autophagy flux. The poly-neddylation dependent autophagic clearance of huntingtin exon1 by HYPK leads to better cell physiology and survival. Taken together, our study describes a novel mechanism of HYPK mediated autophagy of poly-neddylated huntingtin exon1 aggregates.

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Expression, Functional Characterization and X-ray Analysis of HosA, A Member of MarR Family of Transcription Regulator from Uropathogenic Escherichia coli

Cited by 11 publications

References 33 publications

A functional study of the global transcriptional regulator PadR from a strain Streptomyces fradiae‐nitR+bld, resistant to nitrone‐oligomycin

A functional study of the global transcriptional regulator PadR from a strain Streptomyces fradiae‐nitR+bld, resistant to nitrone‐oligomycin

CtcS, a MarR family regulator, regulates chlortetracycline biosynthesis

HYPK scaffolds the Nedd8 and LC3 proteins to initiate formation of autophagosome around polyneddylated huntingtin exon1 aggregates

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