DNA binding properties of the LexA repressor

Schnarr, Manfred; Oertel-Buchheit, Pascale; Kazmaier, Michaël; Granger-Schnarr, Michéle

doi:10.1016/0300-9084(91)90109-e

Cited by 109 publications

(106 citation statements)

References 46 publications

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“…The SOS regulon of E. coli is involved in various cellular processes, such as nucleotide excision and recombination repair (uvrABD, recN, ruvAB), error-prone replication (polB, dinB, umuDC) and inhibition of cell division (sulA). The degree of induction of SOS gene expression depends on at least four parameters: (i) the affinity of LexA for the SOS box, (ii) the location of the SOS box relative to the promoter, (iii) the promoter strength, and (iv) the presence of any additional constitutive promoters (Friedberg et al, 2005;Schnarr et al, 1991;Walker, 1996). Detailed examination of LexA binding in E. coli has been conducted in two studies using genome-wide techniques and chromatin immunoprecipitation (ChIP)-to-chip analysis, enabling the classification of 31 and 49 genes into the LexA regulon, respectively (Fernández de Henestrosa et al, 2000;Wade et al, 2005).…”

Section: Introductionmentioning

confidence: 99%

Genetic makeup of the Corynebacterium glutamicum LexA regulon deduced from comparative transcriptomics and in vitro DNA band shift assays

et al. 2009

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The lexA gene of Corynebacterium glutamicum ATCC 13032 was deleted to create the mutant strain C. glutamicum NJ2114, which has an elongated cell morphology and an increased doubling time. To characterize the SOS regulon in C. glutamicum, the transcriptomes of NJ2114 and a DNA-damage-induced wild-type strain were compared with that of a wild-type control using DNA microarray hybridization. The expression data were combined with bioinformatic pattern searches for LexA binding sites, leading to the detection of 46 potential SOS boxes located upstream of differentially expressed transcription units. Binding of a hexahistidyl-tagged LexA protein to 40 double-stranded oligonucleotides containing the potential SOS boxes was demonstrated in vitro by DNA band shift assays. It turned out that LexA binds not only to SOS boxes in the promoteroperator region of upregulated genes, but also to SOS boxes detected upstream of downregulated genes. These results demonstrated that LexA controls directly the expression of at least 48 SOS genes organized in 36 transcription units. The deduced genes encode a variety of physiological functions, many of them involved in DNA repair and survival after DNA damage, but nearly half of them have hitherto unknown functions. Alignment of the LexA binding sites allowed the corynebacterial SOS box consensus sequence TcGAA(a/c)AnnTGTtCGA to be deduced. Furthermore, the common intergenic region of lexA and the differentially expressed divS-nrdR operon, encoding a cell division suppressor and a regulator of deoxyribonucleotide biosynthesis, was characterized in detail. Promoter mapping revealed differences in divS-nrdR expression during SOS response and normal growth conditions. One of the four LexA binding sites detected in the intergenic region is involved in regulating divS-nrdR transcription, whereas the other sites are apparently used for negative autoregulation of lexA expression.

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Section: Introductionmentioning

confidence: 99%

Genetic makeup of the Corynebacterium glutamicum LexA regulon deduced from comparative transcriptomics and in vitro DNA band shift assays

et al. 2009

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“…The LexA repressor from Escherichia coli is a 202 residue protein that regulates transcription of -20 genes known as SOS genes, which are involved in DNA repair and replication, mutagenesis and cell division [for reviews see Little and Mount (1982), Walker (1984), Little (1991), Schnarr et al (1991) and Schnarr and Granger-Schnarr (1993)]. LexA dimerizes with a rather weak dissociation constant of 50 ,uM (Schnarr et al, 1985 to form an active dimer that binds to a 16 bp palindromic sequence (Wertman and Mount, 1985).…”

Section: Introductionmentioning

confidence: 99%

Solution structure of the LexA repressor DNA binding domain determined by 1H NMR spectroscopy.

et al. 1994

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The structure of the 84 residue DNA binding domain of the Escherichia coli LexA repressor has been determined from NMR data using distance geometry and restrained molecular dynamics. The assignment of the 1H NMR spectrum of the molecule, derived from 2-and 3-D homonuclear experiments, is also reported. A total of 613 non-redundant distance restraints were used to give a final family of 28 structures. The structured region of the molecule consisted of residues 4-69 and yielded a r.m.s. deviation from an average of 0.9 A for backbone and 1.6 A for all heavy atoms. The structure contains three regular a-helices at residues 6-21 (I), 28-35 (1) and 41-52 (HI), and an antiparailel P-sheet at residues 56-58 and 66-68. Helices II and III form a variant helix-turn-helix DNA binding motif, with an unusual one residue insert at residue 38. The topology of the LexA DNA binding domain is found to be the same as for the DNA binding domains of the catabolic activator protein, human histone 5, the HNF-3/fork head protein and the Kluyveromyces lactis heat shock transcription factor.

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“…Nonetheless, we think that the lack of leucine zipper proteins is due to the design of the bait construct. LexA binds to DNA as a homodimer through its N-terminal domain, while dimerization is mediated by its C-terminal domain (52). The use of NRL-ZIP as LexA fusion bait presents two possible outcomes.…”

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The Leucine Zipper of NRL Interacts with the CRX Homeodomain

Mitton

Swain

Chen

et al. 2000

Journal of Biological Chemistry

191

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Photoreceptor-specific expression of rhodopsin is mediated by multiple cis-acting elements in the proximal promoter region. NRL (neural retina leucine zipper) and CRX (cone rod homeobox) proteins bind to the adjacent NRE and Ret-4 sites, respectively, within this region. Although NRL and CRX are each individually able to induce rhodopsin promoter activity, when expressed together they exhibit transcriptional synergy in rhodopsin promoter activation. Using the yeast two-hybrid method and glutathione S-transferase pull-down assays, we demonstrate that the leucine zipper of NRL can physically interact with CRX. Deletion analysis revealed that the CRX homeodomain (CRX-HD) plays an important role in the interaction with the NRL leucine zipper. Although binding with the CRX-HD alone was weak, a strong interaction was detected when flanking regions including the glutamine-rich and the basic regions that follow the HD were included. A reciprocal deletion analysis showed that the leucine zipper of NRL is required for interaction with CRX-HD. Two disease-causing mutations in CRX-HD (R41W and R90W) that exhibit reduced DNA binding and transcriptional synergy also decrease its interaction with NRL. These studies suggest novel possibilities for protein-protein interaction between two conserved DNA-binding motifs and imply that cross-talk among distinct regulatory pathways contributes to the establishment and maintenance of photoreceptor function.Gene activation is a stringently controlled process, involving combinatorial and cooperative action of multiple regulatory proteins with promoter and enhancer DNA elements (1-6). Recent studies, including reconstitution experiments, have suggested that target specificity and transcriptional synergy are achieved by specific and precise interactions among various activator proteins during the assembly of higher order nucleoprotein complexes, including the "enhanceosome" (7-11). The elucidation of these protein-protein and protein-DNA interactions is critical to our understanding of the mechanisms of cell type-and tissue-specific gene expression.Generation of multiple neuronal cell types during retinal development is an evolutionarily conserved biological process, which offers a convenient model system to investigate tissuespecific gene regulation. More than 30 transcription factors representing several classes of DNA-binding proteins are expressed in developing and mature mammalian retina; nevertheless, the precise function of a majority of these proteins remains to be elucidated (12-14). Rhodopsin, the G-proteincoupled light receptor, is expressed specifically in the rod photoreceptors of retina and is a pivotal protein for visual function. Its expression is correlated to rod differentiation and maintained at high levels afterward, throughout life (15). Altered expression of rhodopsin and mutations that affect its function in mature rods result in retinal degeneration (15,16). Regulation of rhodopsin expression is primarily at the level of transcription and is mediated by two distinct...

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DNA binding properties of the LexA repressor

Cited by 109 publications

References 46 publications

Genetic makeup of the Corynebacterium glutamicum LexA regulon deduced from comparative transcriptomics and in vitro DNA band shift assays

Genetic makeup of the Corynebacterium glutamicum LexA regulon deduced from comparative transcriptomics and in vitro DNA band shift assays

Solution structure of the LexA repressor DNA binding domain determined by 1H NMR spectroscopy.

The Leucine Zipper of NRL Interacts with the CRX Homeodomain

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