scholarly journals Mutational Analysis of the fruA Promoter Region Demonstrates that C-Box and 5-Base-Pair Elements Are Important for Expression of an Essential Developmental Gene of Myxococcus xanthus

2004 ◽  
Vol 186 (17) ◽  
pp. 5961-5967 ◽  
Author(s):  
D. Srinivasan ◽  
Lee Kroos
Keyword(s):  
Gene Expression ◽  
Promoter Region ◽  
Mutational Analysis ◽  
Myxococcus Xanthus ◽  
Developmental Gene ◽  
Promoter Regions ◽  
Regulate Gene Expression ◽  
Extracellular Signals ◽  
First Time ◽  
Regulate Gene

ABSTRACT Myxococcus xanthus uses extracellular signals during development to regulate gene expression. C-signaling regulates the expression of many genes induced after 6 h into development. FruA is a protein that is necessary for cells to respond to C-signaling, but expression of the fruA gene does not depend on C-signaling. Yet the fruA promoter region has a C box and a 5-bp element, similar to the promoter regions of several C-signal-dependent genes, where these sequences are crucial. Here, we show that the C box and 5-bp elements are important for expression of fruA, demonstrating for the first time that these sequences play a role in the expression of a gene that does not depend on C-signaling and is required for M. xanthus development.

scholarly journals DNA methylation subpatterns at distinct regulatory regions in human early embryos

Open Biology ◽  
2018 ◽  
Vol 8 (10) ◽  
pp. 180131 ◽  
Author(s):  
Rongsong Luo ◽  
Chunling Bai ◽  
Lei Yang ◽  
Zhong Zheng ◽  
Guanghua Su ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Correlation Analysis ◽  
Promoter Region ◽  
Promoter Regions ◽  
Regulate Gene Expression ◽  
Early Embryos ◽  
Candidate Promoter ◽  
Single Base Resolution ◽  
Genomic Regions

DNA methylation has been investigated for many years, but recent technologies have allowed for single-cell- and single-base-resolution DNA methylation datasets and more accurate assessment of DNA methylation dynamics at the key genomic regions that regulate gene expression in human early embryonic development. In this study, the region from upstream 20 kb to downstream 20 kb of RefSeq gene was selected and divided into 12 distinct regions (up20, up10, up5, up2, 5'UTR, exon, intron, 3'UTR, down2, down5, down10 and down20). The candidate promoter region (TSS ± 2 kb) was further divided into 20 consecutive subregions, which were termed ‘bins’. The DNA methylation dynamics of these regions were systematically analysed along with their effects on gene expression in human early embryos. The dynamic DNA methylation subpatterns at the distinct genomic regions with a focus on promoter regions were mapped. For the 12 distinct genomic regions, up2 and 5'UTR had the lowest DNA methylation levels, and their methylation dynamics were different with other regions. The region 3'UTR had the highest DNA methylation levels, and the correlation analysis with gene expression proved that it was a feature of transcribed genes. For the 20 bins in promoter region, the CpG densities showed a normal distribution pattern, and the trend of the methylated CpG counts was inverse with the DNA methylation levels, especially for the bin 1 (downstream 200 bp of the TSS). Through the correlation analysis between DNA methylation and gene expression, the current study finally revealed that the region bin −4 to 6 (800 bp upstream to 1200 bp downstream of the TSS) was the best candidate for the promoter region in human early embryos, and bin 1 was the putative key regulator of gene activity. This study provided a global and high-resolution view of DNA methylation subpatterns at the distinct genomic regions in human early embryos.

scholarly journals Mutational Analysis of the Myxococcus xanthus Ω4499 Promoter Region Reveals Shared and Unique Properties in Comparison with Other C-Signal-Dependent Promoters

2004 ◽  
Vol 186 (12) ◽  
pp. 3766-3776 ◽  
Author(s):  
Deborah R. Yoder ◽  
Lee Kroos
Keyword(s):  
Base Pair ◽  
Promoter Region ◽  
Promoter Activity ◽  
Mutational Analysis ◽  
Myxococcus Xanthus ◽  
Promoter Regions ◽  
Multicellular Development ◽  
Σ Factor ◽  
Signal Dependence ◽  
Promoter Mutations

ABSTRACT The bacterium Myxococcus xanthus undergoes multicellular development during times of nutritional stress and uses extracellular signals to coordinate cell behavior. C-signal affects gene expression late in development, including that of Ω4499, an operon identified by insertion of Tn5 lac into the M. xanthus chromosome. The Ω4499 promoter region has several sequences in common with those found previously to be important for expression of other C-signal-dependent promoters. To determine if these sequences are important for Ω4499 promoter activity, the effects of mutations on expression of a downstream reporter gene were tested in M. xanthus. Although the promoter resembles those recognized by Escherichia coli σ54, mutational analysis implied that a σ70-type σ factor likely recognizes the promoter. A 7-bp sequence known as a C box and a 5-bp element located 6 bp upstream of the C box have been shown to be important for expression of other C-signal-dependent promoters. The Ω4499 promoter region has C boxes centered at −33 and −55 bp, with 5-bp elements located 7 and 8 bp upstream, respectively. A multiple-base-pair mutation in any of these sequences reduced Ω4499 promoter activity more than twofold. Single base-pair mutations in the C box centered at −33 bp yielded a different pattern of effects on expression than similar mutations in other C boxes, indicating that each functions somewhat differently. An element from about −81 to −77 bp exerted a twofold positive effect on expression but did not appear to be responsible for the C-signal dependence of the Ω4499 promoter. Mutations in sigD and sigE, which are genes that encode σ factors, reduced expression from the Ω4499 promoter. The results provide further insight into the regulation of C-signal-dependent genes, demonstrating both shared and unique properties among the promoter regions so far examined.

scholarly journals Mutational Analysis of the Myxococcus xanthus Ω4400 Promoter Region Provides Insight into Developmental Gene Regulation by C Signaling

2004 ◽  
Vol 186 (3) ◽  
pp. 661-671 ◽  
Author(s):  
Deborah R. Yoder ◽  
Lee Kroos
Keyword(s):  
Dna Sequences ◽  
Promoter Region ◽  
Mutational Analysis ◽  
Myxococcus Xanthus ◽  
Regulatory Region ◽  
Cell Movement ◽  
Site Directed Mutagenesis ◽  
Promoter Regions ◽  
Regulatory Regions ◽  
The Individual

ABSTRACT Myxococcus xanthus utilizes extracellular signals during development to coordinate cell movement, differentiation, and changes in gene expression. One of these signals, the C signal, regulates the expression of many genes, including Ω4400, a gene identified by an insertion of Tn5 lac into the chromosome. Expression of Tn5 lac Ω4400 is reduced in csgA mutant cells, which fail to perform C signaling, and the promoter region has several sequences similar to sequences found in the regulatory regions of other C-signal-dependent genes. One such gene, Ω4403, depends absolutely on the C signal for expression, and its promoter region has been characterized previously by mutational analysis. To determine if the similar sequences within the Ω4400 and Ω4403 regulatory regions function in the same way, deletion analysis and site-directed mutagenesis of the Ω4400 promoter region were performed. A 7-bp sequence centered at −49 bp, termed a C box, is identical in the Ω4400 and Ω4403 promoter regions, yet mutations in the individual base pairs affected expression from the two promoters very differently. Also, a single-base-pair change within a similar 5-bp element, which is centered at −61 bp in both promoter regions, had very different effects on the activities of the two promoters. Further mutational analysis showed that two regions are important for Ω4400 expression; one region, from −63 to −31 bp, is required for Ω4400 expression, and the other, from −86 to −81 bp, exerts a two- to fourfold effect on expression and is at least partially responsible for the C signal dependence of the Ω4400 promoter. Mutations in sigD and sigE, which are genes that encode σ factors, abolished and reduced Ω4400 expression, respectively. Expression of Ω4400 in actB or actC mutants correlated well with the altered levels of C signal produced in these mutants. Our results provide the first detailed analysis of an M. xanthus regulatory region that depends partially on C signaling for expression and indicate that similar DNA sequences in the Ω4400 and Ω4403 promoter regions function differently.

scholarly journals Optimization of a bacterial three-hybrid assay through in vivo titration of an RNA-DNA adapter-protein

2020 ◽  
Author(s):  
Clara D. Wang ◽  
Rachel Mansky ◽  
Hannah LeBlanc ◽  
Chandra M. Gravel ◽  
Katherine E. Berry
Keyword(s):  
Gene Expression ◽  
Protein Interactions ◽  
Regulate Gene Expression ◽  
E Coli ◽  
Binding Energetics ◽  
Response To Stress ◽  
First Time ◽  
Regulate Gene

ABSTRACTNon-coding RNAs regulate gene expression in every domain of life. In bacteria, small RNAs (sRNAs) regulate gene expression in response to stress and are often assisted by RNA-chaperone proteins, such as Hfq. We have recently developed a bacterial three-hybrid (B3H) assay that detects the strong binding interactions of certain E. coli sRNAs with proteins Hfq and ProQ. Despite the promise of this system, the signal-to-noise has made it challenging to detect weaker interactions. In this work, we use Hfq-sRNA interactions as a model system to optimize the B3H assay, so that weaker RNA-protein interactions can be more reliably detected. We find that the concentration of the RNA-DNA adapter is an important parameter in determining the signal in the system, and have modified the plasmid expressing this component to tune its concentration to optimal levels. In addition, we have systematically perturbed the binding affinity of Hfq-RNA interactions to define, for the first time, the relationship between B3H signal and in vitro binding energetics. The new pAdapter construct presented here substantially expands the range of detectable interactions in the B3H assay, broadening its utility. This improved assay will increase the likelihood of identifying novel protein-RNA interactions with the B3H system, and will facilitate exploration of the binding mechanisms of these interactions.

Human cytomegalovirus US3 and UL36-38 immediate-early proteins regulate gene expression.

1992 ◽  
Vol 66 (1) ◽  
pp. 95-105 ◽  
Author(s):  
A M Colberg-Poley ◽  
L D Santomenna ◽  
P P Harlow ◽  
P A Benfield ◽  
D J Tenney
Keyword(s):  
Gene Expression ◽  
Human Cytomegalovirus ◽  
Immediate Early ◽  
Regulate Gene Expression ◽  
Early Proteins ◽  
Immediate Early Proteins ◽  
Regulate Gene

scholarly journals Arabidopsis heat shock transcription factor HSFA7b positively mediates salt stress tolerance by binding to an E-box-like motif to regulate gene expression

2019 ◽  
Vol 70 (19) ◽  
pp. 5355-5374 ◽  
Author(s):  
Dandan Zang ◽  
Jingxin Wang ◽  
Xin Zhang ◽  
Zhujun Liu ◽  
Yucheng Wang
Keyword(s):  
Gene Expression ◽  
Transcription Factors ◽  
Heat Shock ◽  
Salt Tolerance ◽  
Stress Responses ◽  
Ion Homeostasis ◽  
Cellulose Synthase ◽  
Regulate Gene Expression ◽  
E Box ◽  
Regulate Gene

Abstract Plant heat shock transcription factors (HSFs) are involved in heat and other abiotic stress responses. However, their functions in salt tolerance are little known. In this study, we characterized the function of a HSF from Arabidopsis, AtHSFA7b, in salt tolerance. AtHSFA7b is a nuclear protein with transactivation activity. ChIP-seq combined with an RNA-seq assay indicated that AtHSFA7b preferentially binds to a novel cis-acting element, termed the E-box-like motif, to regulate gene expression; it also binds to the heat shock element motif. Under salt conditions, AtHSFA7b regulates its target genes to mediate serial physiological changes, including maintaining cellular ion homeostasis, reducing water loss rate, decreasing reactive oxygen species accumulation, and adjusting osmotic potential, which ultimately leads to improved salt tolerance. Additionally, most cellulose synthase-like (CSL) and cellulose synthase (CESA) family genes were inhibited by AtHSFA7b; some of them were randomly selected for salt tolerance characterization, and they were mainly found to negatively modulate salt tolerance. By contrast, some transcription factors (TFs) were induced by AtHSFA7b; among them, we randomly identified six TFs that positively regulate salt tolerance. Thus, AtHSFA7b serves as a transactivator that positively mediates salinity tolerance mainly through binding to the E-box-like motif to regulate gene expression.

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