A software program combining sequence motif searches with keywords for finding repeats containing DNA sequences

Structural differentiation of bacterial chromatin depends on cooperative binding of abundant nucleoid-associated proteins at numerous genomic DNA sites and stabilization of distinct long-range nucleoprotein structures. Histone-like nucleoid-structuring protein (H-NS) is an abundant DNA-bridging, nucleoid-associated protein that binds to an AT-rich conserved DNA sequence motif and regulates both the shape and the genetic expression of the bacterial chromosome. Although there is ample evidence that the mode of H-NS binding depends on environmental conditions, the role of the spatial organization of H-NS-binding sequences in the assembly of long-range nucleoprotein structures remains unknown. In this study, by using high-resolution atomic force microscopy combined with biochemical assays, we explored the formation of H-NS nucleoprotein complexes on circular DNA molecules having different arrangements of identical sequences containing high-affinity H-NS-binding sites. We provide the first experimental evidence that variable sequence arrangements result in various three-dimensional nucleoprotein structures that differ in their shape and the capacity to constrain supercoils and compact the DNA. We believe that the DNA sequence-directed versatile assembly of periodic higher-order structures reveals a general organizational principle that can be exploited for knowledge-based design of long-range nucleoprotein complexes and purposeful manipulation of the bacterial chromatin architecture.

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Binding sites of the 9- and 14-kilodalton heterodimeric protein subunit of the signal recognition particle (SRP) are contained exclusively in the Alu domain of SRP RNA and contain a sequence motif that is conserved in evolution.

Molecular and Cellular Biology ◽

10.1128/mcb.11.8.3949 ◽

1991 ◽

Vol 11 (8) ◽

pp. 3949-3959 ◽

Cited By ~ 56

Author(s):

K Strub ◽

J Moss ◽

P Walter

Keyword(s):

Dna Sequences ◽

Close Contact ◽

Signal Recognition Particle ◽

Protein Translation ◽

Endoplasmic Reticulum Membrane ◽

Sequence Motif ◽

Signal Recognition ◽

Protein Subunit ◽

Rnase Protection ◽

Srp Rna

The mammalian signal recognition particle (SRP) is a small cytoplasmic ribonucleoprotein required for the cotranslational targeting of secretory proteins to the endoplasmic reticulum membrane. The heterodimeric protein subunit SRP9/14 was previously shown to be essential for SRP to cause pausing in the elongation of secretory protein translation. RNase protection and filter binding experiments have shown that binding of SRP9/14 to SRP RNA depends solely on sequences located in a domain of SRP RNA that is strongly homologous to the Alu family of repetitive DNA sequences. In addition, the use of hydroxyl radicals, as RNA-cleaving reagents, has revealed four distinct regions in this domain that are in close contact with SRP9/14. Surprisingly, the nucleotide sequence in one of these contact sites, predicted to be mostly single stranded, was found to be extremely conserved in SRP RNAs of evolutionarily distant organisms ranging from eubacteria and archaebacteria to yeasts and higher eucaryotic cells. This finding suggests that SRP9/14 homologs may also exist in these organisms, where they possibly contribute to the regulation of protein synthesis similar to that observed for mammalian SRP in vitro.

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Novel structural organisation of a Mus musculus DBA/2 chromosome shows a fixed position for the centromere

Journal of Cell Science ◽

10.1242/jcs.106.1.79 ◽

1993 ◽

Vol 106 (1) ◽

pp. 79-85 ◽

Cited By ~ 3

Author(s):

A.R. Mitchell ◽

L. Nicol ◽

P. Malloy ◽

D. Kipling

Keyword(s):

Dna Sequences ◽

Inbred Mouse ◽

Inbred Mouse Strain ◽

Chromosome 1 ◽

Sequence Motif ◽

Major Satellite ◽

Satellite Sequences ◽

Minor Satellite ◽

Apparent Similarity ◽

Terminal Block

Chromosome 1 of the inbred mouse strain DBA/2 shows an unusual polymorphism associated with its centromeric satellite DNA sequences. The minor satellite array has undergone amplification and is present as two blocks separated by major satellite sequences. Both minor satellite blocks appear to carry the sequence motif necessary for CENP-B protein binding. Despite this apparent similarity the functional centromere, as defined by the location of CREST antigens, appears to form only within the more terminal block. The two blocks also vary in that sister chromatid association only occurs with this more terminal block.

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Nonhomologous recombination in the parvovirus chromosome: role for a CTATTTCT motif

Molecular and Cellular Biology ◽

10.1128/mcb.6.8.3005-3009.1986 ◽

1986 ◽

Vol 6 (8) ◽

pp. 3005-3009

Author(s):

A Hogan ◽

E A Faust

Keyword(s):

Dna Sequences ◽

Topoisomerase I ◽

Sequence Motif ◽

Cleavage Sites ◽

Genetic Element ◽

Minute Virus Of Mice ◽

Nonhomologous Recombination ◽

Naturally Occurring ◽

Minute Virus ◽

Foreign Dna

The mechanism of nonhomologous recombination in murine cells infected with the parvovirus minute virus of mice (MVM) has been investigated by analysis of DNA sequences at recombination junctions in naturally occurring deletion variants of the virus. We report here that nonhomologous recombination in the MVM chromosome is characterized by short homologies, by insertion at recombination junctions of foreign DNA sequences that are enriched for preferred eucaryotic topoisomerase I cleavage sites, and by an association with a common DNA sequence motif of the type 5'-CTATTTCT-3'. Additional analyses of broken MVM chromosomes provided evidence for specific enzymatic cleavage within 5'-CTTATC-3' and 5'-CTATTC-3' sequences. The results indicate that the 5'-CTATTTCT-3' motif is an important genetic element for nonhomologous recombination in the parvovirus chromosome.

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The Purine Repressor of Bacillus subtilis: a Novel Combination of Domains Adapted for Transcription Regulation

Journal of Bacteriology ◽

10.1128/jb.185.14.4087-4098.2003 ◽

2003 ◽

Vol 185 (14) ◽

pp. 4087-4098 ◽

Cited By ~ 23

Author(s):

Sangita C. Sinha ◽

Joseph Krahn ◽

Byung Sik Shin ◽

Diana R. Tomchick ◽

Howard Zalkin ◽

...

Keyword(s):

Bacillus Subtilis ◽

Dna Binding ◽

Dna Sequences ◽

Sequence Motif ◽

Charged Surface ◽

Winged Helix ◽

Gram Positive Bacteria ◽

Terminal Domain ◽

Winged Helix Domain ◽

Positively Charged

ABSTRACT The purine repressor from Bacillus subtilis, PurR, represses transcription from a number of genes with functions in the synthesis, transport, and metabolism of purines. The 2.2-Å crystal structure of PurR reveals a two-domain protein organized as a dimer. The larger C-terminal domain belongs to the PRT structural family, in accord with a sequence motif for binding the inducer phosphoribosylpyrophosphate (PRPP). The PRT domain is fused to a smaller N-terminal domain that belongs to the winged-helix family of DNA binding proteins. A positively charged surface on the winged-helix domain likely binds specific DNA sequences in the recognition site. A second positively charged surface surrounds the PRPP site at the opposite end of the PurR dimer. Conserved amino acids in the sequences of PurR homologs in 21 gram-positive bacteria cluster on the proposed recognition surface of the winged-helix domain and around the PRPP binding site at the opposite end of the molecule, supporting a common function of DNA and PRPP binding for all of the proteins. The structure supports a binding mechanism in which extended regions of DNA interact with extensive protein surface. Unlike most PRT proteins, which are phosphoribosyltransferases (PRTases), PurR lacks catalytic activity. This is explained by a tyrosine side chain that blocks the site for a nucleophile cosubstrate in PRTases. Thus, B. subtilis has adapted an enzyme fold to serve as an effector-binding domain and has used it in a novel combination with the DNA-binding winged-helix domain as a repressor of purine genes.

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Insights into DNA substrate selection by APOBEC3G from structural, biochemical, and functional studies

10.1101/235481 ◽

2017 ◽

Author(s):

Samantha J. Ziegler ◽

Chang Liu ◽

Mark Landau ◽

Olga Buzovetsky ◽

Belete A. Desimmie ◽

...

Keyword(s):

Dna Sequences ◽

Binding Pocket ◽

Sequence Motif ◽

Specific Substrate ◽

Substrate Selection ◽

Cytidine Deaminases ◽

Human Apolipoprotein ◽

Functional Studies ◽

Dna Substrate ◽

Hiv 1

AbstractHuman apolipoprotein B mRNA-editing enzyme-catalytic polypeptide-like 3 (A3) proteins are a family of cytidine deaminases that catalyze the conversion of cytidine to uridine in single-stranded DNA (ssDNA). A3 proteins act in the innate immune response to viral infection by mutating the viral ssDNA. One of the most well-studied human A3 family members is A3G, which is a potent inhibitor of HIV-1. Each A3 protein prefers a specific substrate sequence for catalysis - for example, A3G deaminates the third cytidine in the CCCA sequence motif. However, the interaction between A3G and ssDNA is difficult to characterize due to poor solution behavior of the full-length protein and loss of DNA affinity of the truncated protein. Here, we present a novel DNA-anchoring fusion strategy, which we have used to capture an A3G-ssDNA interaction. We characterized an A3G-DNA binding pocket that is important for the enzyme to scan the DNA for its hotspot. The results provide insights into the mechanism by which A3G selects and deaminates its preferred substrates and help define how A3 proteins are tailored to recognize specific DNA sequences. This knowledge contributes to a better understanding of the mechanism of DNA substrate selection by A3G, as well as A3G antiviral activity against HIV-1.

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Localization of transcriptional regulatory elements and nuclear factor binding sites in mouse ribosomal protein gene rpL32

Molecular and Cellular Biology ◽

10.1128/mcb.9.5.2067-2074.1989 ◽

1989 ◽

Vol 9 (5) ◽

pp. 2067-2074

Author(s):

M L Atchison ◽

O Meyuhas ◽

R P Perry

Keyword(s):

Ribosomal Protein ◽

Dna Sequences ◽

Binding Sites ◽

Protein Gene ◽

Transcriptional Start Site ◽

Ribosomal Protein Gene ◽

Regulatory Elements ◽

Sequence Motif ◽

Start Site ◽

Nuclear Factors

The DNA sequences required for expression of the ribosomal protein gene rpL32 were identified by transient-expression assays of chimeric rpL32-chloramphenicol acetyltransferase genes. These studies showed that maximal rpL32 expression requires sequences in a 150- to 200-base-pair region spanning the transcriptional start site. Three discrete regions of importance were identified: one between positions -79 and -69 and two others located downstream of the transcriptional start site. Progressive 5' or 3' deletions caused stepwise decreases in expression, which suggested a complex interplay of redundant or compensatory elements. Gel mobility shift assays were used to identify trans-acting nuclear factors which bind to segments of the rpL32 promoter that are known to be important for transcription. Evidence for several distinct nuclear factors is presented. The binding sites for these factors were localized to the following regions: -79 to -69, -36 to -19, -19 to +11, +11 to +46 in exon I, and within the first 31 base pairs of intron 1. One of these factors may bind to multiple sites within the promoter region. Interestingly, the factor that binds to a sequence motif in the first exon also binds to similar motifs in a comparable region of the c-myc gene.

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Localization of transcriptional regulatory elements and nuclear factor binding sites in mouse ribosomal protein gene rpL32.

Molecular and Cellular Biology ◽

10.1128/mcb.9.5.2067 ◽

1989 ◽

Vol 9 (5) ◽

pp. 2067-2074 ◽

Cited By ~ 43

Author(s):

M L Atchison ◽

O Meyuhas ◽

R P Perry

Keyword(s):

Ribosomal Protein ◽

Dna Sequences ◽

Binding Sites ◽

Protein Gene ◽

Transcriptional Start Site ◽

Ribosomal Protein Gene ◽

Regulatory Elements ◽

Sequence Motif ◽

Start Site ◽

Nuclear Factors

The DNA sequences required for expression of the ribosomal protein gene rpL32 were identified by transient-expression assays of chimeric rpL32-chloramphenicol acetyltransferase genes. These studies showed that maximal rpL32 expression requires sequences in a 150- to 200-base-pair region spanning the transcriptional start site. Three discrete regions of importance were identified: one between positions -79 and -69 and two others located downstream of the transcriptional start site. Progressive 5' or 3' deletions caused stepwise decreases in expression, which suggested a complex interplay of redundant or compensatory elements. Gel mobility shift assays were used to identify trans-acting nuclear factors which bind to segments of the rpL32 promoter that are known to be important for transcription. Evidence for several distinct nuclear factors is presented. The binding sites for these factors were localized to the following regions: -79 to -69, -36 to -19, -19 to +11, +11 to +46 in exon I, and within the first 31 base pairs of intron 1. One of these factors may bind to multiple sites within the promoter region. Interestingly, the factor that binds to a sequence motif in the first exon also binds to similar motifs in a comparable region of the c-myc gene.

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Characterization of the Plasmid Encoded Virulence Region pat-1 of Phytopathogenic Clavibacter michiganensis subsp. michiganensis

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi.1997.10.2.195 ◽

1997 ◽

Vol 10 (2) ◽

pp. 195-206 ◽

Cited By ~ 69

Author(s):

Jens Dreier ◽

Dietmar Meletzus ◽

Rudolf Eichenlaub

Keyword(s):

Structural Gene ◽

Dna Sequences ◽

Repetitive Sequence ◽

Transcription Initiation ◽

Serine Proteases ◽

Tandem Repeats ◽

Sequence Motif ◽

Clavibacter Michiganensis ◽

Reading Frame ◽

Clavibacter Michiganensis Subsp

The tomato pathogen Clavibacter michiganensis subsp. michiganensis NCPPB382, causing bacterial wilt and canker, harbors two plasmids, pCM1 (27.5 kb) and pCM2 (72 kb), carrying genes involved in virulence. The region of plasmid pCM2 encoding the pathogenicity locus pat-1 was mapped by deletion analysis and complementation studies to a 1.5-kb BglII/SmaI DNA fragment. Introduction of the pat-1 region into endophytic, plasmid-free isolates of C. michiganensis subsp. michiganensis converted these bacteria into virulent pathogens. Based on the nucleotide sequence of the pat-1 region, an open reading frame (ORF1) can be predicted, coding for a protein of 280 amino acids and 29.7 kDa with homology to serine proteases. Introduction of a frame-shift mutation in ORF1 leads to a loss of the pathogenic phenotype. Northern (RNA) hybridizations identified an 1.5-knt transcript of the pat-1 structural gene. The site of transcription initiation was mapped by primer extension and a typical -10/-35 region was located with significant homology to the consensus Escherichia coli σ70 and Bacillus subtilis σ43 promoters. Downstream of the pat-1 structural gene, a peculiar repetitive sequence motif (pat-1rep) is located, consisting of 20 direct tandem repeats preceded by a run of 14 guanosine residues. DNA sequences homologous to pat-1rep were isolated and characterized from four virulent C. michiganensis subsp. mich-iganensis strains exhibiting a high extent of structural conservation. The deletion of this repetitive sequence reduced virulence significantly but did not lead to a complete loss of the virulence phenotype.

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Role of Transcription Factor Kar4 in Regulating Downstream Events in the Saccharomyces cerevisiae Pheromone Response Pathway

Molecular and Cellular Biology ◽

10.1128/mcb.00439-06 ◽

2006 ◽

Vol 27 (3) ◽

pp. 818-829 ◽

Cited By ~ 26

Author(s):

Ron Lahav ◽

Alison Gammie ◽

Saeed Tavazoie ◽

Mark D. Rose

Keyword(s):

Transcription Factor ◽

Dna Sequences ◽

Critical Role ◽

Gene Clusters ◽

Sequence Motif ◽

Dependent Manner ◽

Regulatory Dna Sequences ◽

Putative Transcription Factor ◽

Regulatory Dna

ABSTRACT Yeast Kar4 is a putative transcription factor required for karyogamy (the fusion of haploid nuclei during mating) and possibly other functions. Previously known to be required only for the transcriptional induction of KAR3 and CIK1, microarray experiments identified many genes regulated by Kar4 in both mating and mitosis. Several gene clusters are positively or negatively regulated by mating pheromone in a Kar4-dependent manner. Chromatin immunoprecipitation and gel shift assays confirmed that Kar4 binds to regulatory DNA sequences upstream of KAR3. Together with one-hybrid experiments, these data support a model in which both Kar4 and Ste12 bind jointly to the KAR3 promoter. Analysis of the upstream regions of Kar4-induced genes identified a DNA sequence motif that may be a binding site for Kar4. Mutation within the motif upstream of KAR3 eliminated pheromone induction. Genes regulated by Kar4, on average, are delayed in their temporal expression and exhibit a more stringent dose response to pheromone. Furthermore, the induction of Kar4 by pheromone is necessary for the delayed temporal induction of KAR3 and PRM2, genes required for efficient nuclear fusion during mating. Accordingly, we propose that Kar4 plays a critical role in the choreography of the mating response.

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