Purification and characterization of the Ner repressor of bacteriophage Mu

Kukolj, George; Tolias, Peter; DuBow, Michael S.

doi:10.1016/0014-5793(89)80565-4

Cited by 9 publications

(5 citation statements)

References 28 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…D108 repressor did not bind to any other sites in the intergenic region. D108's Pc promoter is situated to the right of the Ner-operator, as is Mu's (11,20,21) and a corresponding 03 in D108, overlapping this RNA polymerase binding site, was not detected. How D108 regulates transcription from its Pc promoter and the manner in which lysogeny and stable c expression is maintained remains to be determined.…”

Section: Discussionmentioning

confidence: 91%

“…The heterologous phages each encode two negative regulatory proteins of transcription, the products of genes c and ner, which bind to specific sites in their respective intergenic regions (15)(16)(17)(18)(19)(20)(21). Ner is functionally analogous to X Cro in that it negatively regulates transcription from Pc and autoregulates Pe activity (22)(23)(24)(25).…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Characterization of the lysogenic repressor(c)from transposable Mu-like bacteriophage D108

Kukolj

DuBow

1991

Nucl Acids Res

Self Cite

View full text Add to dashboard Cite

The c gene products from related, transposable phages Mu and D108 encode lysogenic repressors which negatively regulate transcription and transposition. Using the gel shift assay to monitor c-operator specific DNA-binding activity, the 19.5 kDa D108 c repressor was purified to homogeneity. Sequence analysis of the N-terminus confirmed the identity of the purified protein as the repressor and ascribed its ATG initiation codon to base pair 864 from the D108 left end. Analytical gel filtration and dimethyl suberimidate cross-linking of repressor at 0.1-0.5 microM concentrations revealed that the repressor protein could form oligomers in the absence of its DNA substrate. From DNase I footprinting and gel mobility shift analyses, the D108 repressor only bound to two operators (O1 and O2) which, as in Mu, flank an Integration Host Factor (IHF) binding site. In contrast to Mu, an O3 site in D108 was not found. Moreover, D108 repressor first bound operator O2, while occupancy of O1 required higher protein concentrations. The implications of these results on the D108 regulatory system are discussed.

show abstract

Section: Discussionmentioning

confidence: 91%

Section: Introductionmentioning

confidence: 99%

Characterization of the lysogenic repressor(c)from transposable Mu-like bacteriophage D108

Kukolj

DuBow

1991

Nucl Acids Res

Self Cite

View full text Add to dashboard Cite

show abstract

“…Ner was cloned into NcoI and BamHI sites of pET28 to generate pWY72, which was transformed into BL21(DE3)pLys E. coli host. Ner expression was induced with 1 mM IPTG at 37C, and purified as described (Kukolj et al , 1989). …”

Section: Methodsmentioning

confidence: 99%

Controlling DNA degradation from a distance: a new role for the Mu transposition enhancer

Choi

Saha

Jang

et al. 2014

Molecular Microbiology

View full text Add to dashboard Cite

Summary Phage Mu is unique among transposable elements in employing a transposition enhancer. The enhancer DNA segment is the site where the transposase MuA binds and makes bridging interactions with the two Mu ends, interwrapping the ends with the enhancer in a complex topology essential for assembling a catalytically active transpososome. The enhancer is also the site at which regulatory proteins control divergent transcription of genes that determine the phage lysis-lysogeny decision. Here we report a third function for the enhancer - that of regulating degradation of extraneous DNA attached to both ends of infecting Mu. This DNA is protected from nucleases by a phage protein until Mu integrates into the host chromosome, after which it is rapidly degraded. We find that leftward transcription at the enhancer, expected to disrupt its topology within the transpososome, blocks degradation of this DNA. Disruption of the enhancer would lead to the loss or dislocation of two non-catalytic MuA subunits positioned in the transpososome by the enhancer. We provide several lines of support for this inference, and conclude that these subunits are important for activating degradation of the flanking DNA. This work also reveals a role for enhancer topology in phage development.

show abstract

“…15.3 ; our unpublished data). Notably, the two ORFs located upstream of alpI share homology with the phage genes ogr and ner ; Ogr is described as a positive regulatory factor for phage P2 late gene transcription while Ner is a negative regulator of Mu bacteriophage early gene transcription (Birkeland et al 1991 ;Kukolj et al 1989 ). The downstream region of alpR reveals several ORFs whose products share similarity with tail proteins of bacteriophage T2 particles and notably with the FETUD operon encoding the major structural components of the P2 contractile tail (Christie et al 2002 ).…”

Section: Genome Analysesmentioning

confidence: 98%

Cell–Cell Communication in Azospirillum and Related PGPR

Wisniewski‐Dyé¹,

Vial²

2015

Handbook for Azospirillum

View full text Add to dashboard Cite

Quorum-sensing (QS) regulation based on N -acyl-L -homoserine lactones (AHL) is used to control various phenotypes that are often essential for interaction with a eukaryotic host, notably for plant-associated bacteria. Technical methodologies currently used to reveal AHL production in a specifi c strain and to decipher phenotypes under QS regulation are surveyed in this chapter. Analyses conducted on the genus Azospirillum and on other related PGPR are used to illustrate the different steps of the approach. Among the genus Azospirillum , a survey of 40 strains belonging to six species revealed AHL production for four strains belonging to the lipoferum species or to a close undefi ned species and isolated from a rice rhizosphere. Identifi cation of genes mediating QS and regulating functions indicate that (1) distinct QS networks are present in some strains and seem to have been acquired independently by horizontal gene transfer; (2) QS regulation is strain specifi c with several phenotypes and numerous proteins being regulated by AHL-based QS in A. lipoferum B518, whereas no change is observed in A. lipoferum TVV3 defi cient in AHL production; and (3) QS is dedicated to regulating functions linked to rhizosphere competence and adaptation to plant roots in A. lipoferum B518.

show abstract

Purification and characterization of the Ner repressor of bacteriophage Mu

Cited by 9 publications

References 28 publications

Characterization of the lysogenic repressor(c)from transposable Mu-like bacteriophage D108

Characterization of the lysogenic repressor(c)from transposable Mu-like bacteriophage D108

Controlling DNA degradation from a distance: a new role for the Mu transposition enhancer

Cell–Cell Communication in Azospirillum and Related PGPR

Contact Info

Product

Resources

About