Download or read book Regulation of the DnaA Promoter in Escherichia Coli written by Victoria Goehner Newman and published by . This book was released on 1999 with total page 492 pages. Available in PDF, EPUB and Kindle. Book excerpt: The timing of initiation of DNA replication from oriC in E. coli is regulated by the concentration of the DnaA protein. Increases in the intracellular DnaA protein concentration over 1.5 fold normal levels causes asynchronous timing of initiation of DNA replication. The regulators of the dnaA operon must keep the DnaA concentration within tight limits yet be able to respond to rapidly changing environmental conditions. Here it is shown that Fis and DnaA proteins interact, in conjunction with other factors, to regulate the expression of dnaA. Binding of DnaA to the DnaA box in the dnaA promoter region is destabilized by the binding of Fis to its binding site in dnaAp2. However, DnaA94, containing only the C-terminal 94 amino acids of the DnaA protein and retaining specific binding activity to the DnaA box of the dnaA promoter region, is not destabilized by Fis binding. DNase I and copper-phenanthroline (Cu-op) footprinting studies of DnaA and DnaA94 support the existence of a second weak DnaA box which abuts the DNase I and Cu-op footprints of Fis in the dnaAp2 promoter. We determined that the GC-rich region of the dnaAp2 promoter is analogous to the discriminator sequence (DS) of rRNA promoters, in that the GC-rich sequence mediates the repression caused by elevated levels of guanosine tetraphosphate (ppGpp). During starvation conditions, the alarmone ppGpp is synthesized. Overexpression of ppGpp causes inhibition of dnaAp2 promoter transcription, which is alleviated when the GC-rich DS is mutated to an AT-rich sequence. We conclude that this site is responsible for mediating the stringent control of the dnaAp2 promoter. Incorrect timing of initiation in the cell cycle results in asynchronous replication. Strains with a mutated DS, DnaA box, or Fis binding site in the dnaA promoter region on the chromosome have increased levels of dnaA gene expression. These mutants initiate DNA replication synchronously in minimal M9 medium as shown by flow cytometric assays. In rich medium, the DS, mutant showed very limited asynchrony, whereas the other mutants replicated synchronously.

Download or read book Growth Rate Regulation and Control of Initiation of DNA Replication in Escherichia Coli written by Anne Eliane Chiaramello and published by . This book was released on 1990 with total page 300 pages. Available in PDF, EPUB and Kindle. Book excerpt: The initiation of DNA synthesis at the chromosomal replication origin, oriC, in Escherichia coli involves an RNA polymerase-mediated step. The level of synthesis of transcripts moving counterclockwise toward oriC is controlled at two promoters, P1 (asnC) and P2 (mioC), and at two transcription terminator regions, T1 and T2. As shown by S1 mapping, termination at the T2 region occurs to the right of oriC at nucleotides 297-299 and 306-310, while major termination events in the T1 region occur in and near the mioC promoter. The majority of transcripts entering oriC originates from the mioC promoter. Transcription from the mioC promoter has been shown to enhance the frequency of initiation of DNA replication of oriC containing plasmids, and to stabilize these plasmids in the host cells. The mioC promoter, which is stringently controlled, is also growth rate regulated. The amount of mioC transcripts relative to the amount of total RNA was inversely correlated with growth rate. This transcript is characterized by a short half-life (1.5 min). The mioC promoter, which contains a DnaA protein binding site, was much less susceptible to repression by DnaA protein when located in the chromosome, than when located in a plasmid. Only a very high concentration of DnaA protein repressed the mioC promoter. The DnaA protein, which is required for initiation of DNA replication from oriC, is growth rate regulated. As shown by RNase protection, this regulation is exerted at the transcriptional level, affecting both promoters, dnaAp1 and dnaAp2. Transcription from these two promoters is also stringently controlled. The amount of DnaA protein in spoT mutants, which are deficient in ppGpp pyrophosphorylase activity, decreases as the severity in the mutation increases. Thus, the intracellular concentration of ppGpp influences the expression of the dnaA gene. In conclusion, the growth rate regulation and stringent control of the dnaA gene suggest that one way in which DNA replication is coordinated with the growth rate is via ppGpp synthesis at the ribosome.

Download or read book Regulation of Gene Expression in Escherichia coli written by E. C. C. Lin and published by Springer Science & Business Media. This book was released on 2012-12-06 with total page 1010 pages. Available in PDF, EPUB and Kindle. Book excerpt: This up-to-date guide focuses on the understanding of key regulatory mechanisms governing gene expression in Escherichia coli. Studies of E. coli not only provide the first models of gene regulation, but research continues to yield different control mechanisms.

Download or read book Identification of Genes Regulated by DNAA Protein in Escherichia Coli written by Qingping Wang and published by . This book was released on 1989 with total page 442 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Growth Rate Regulation and Control of Initiation of DNA Replication in E s c h e r i c h i a C o l i written by and published by . This book was released on 1990 with total page 260 pages. Available in PDF, EPUB and Kindle. Book excerpt: The initiation of DNA synthesis at the chromosomal replication origin, oriC, in Escherichia coli involves an RNA polymerase-mediated step. The level of synthesis of transcripts moving counterclockwise toward oriC is controlled at two promoters, P1 (asnC) and P2 (mioC), and at two transcription terminator regions, T1 and T2. As shown by S1 mapping, termination at the T2 region occurs to the right of oriC at nucleotides 297-299 and 306-310, while major termination events in the T1 region occur in and near the mioC promoter. The majority of transcripts entering oriC originates from the mioC promoter. Transcription from the mioC promoter has been shown to enhance the frequency of initiation of DNA replication of oriC containing plasmids, and to stabilize these plasmids in the host cells. The mioC promoter, which is stringently controlled, is also growth rate regulated. The amount of mioC transcripts relative to the amount of total RNA was inversely correlated with growth rate. This transcript is characterized by a short half-life (1.5 min). The mioC promoter, which contains a DnaA protein binding site, was much less susceptible to repression by DnaA protein when located in the chromosome, than when located in a plasmid. Only a very high concentration of DnaA protein repressed the mioC promoter. The DnaA protein, which is required for initiation of DNA replication from oriC, is growth rate regulated. As shown by RNase protection, this regulation is exerted at the transcriptional level, affecting both promoters, dnaAp1 and dnaAp2. Transcription from these two promoters is also stringently controlled. The amount of DnaA protein in spoT mutants, which are deficient in ppGpp pyrophosphorylase activity, decreases as the severity in the mutation increases. Thus, the intracellular concentration of ppGpp influences the expression of the dnaA gene. In conclusion, the growth rate regulation and stringent control of the dnaA gene suggest that one way in which DNA replication is coordinated with the growth rate is via ppGpp synthesis at the ribosome.

Download or read book Regulation of Chromosomal DNA Replication in Escherichia Coli written by Lyle Adair Simmons and published by . This book was released on 2003 with total page 464 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Analysis of the Stability of Escherichia Coli DnaA Protein in the Regulation of DNA Replication written by Marta Halszka Gross and published by . This book was released on 2018 with total page pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Regulation of Ribonucleotide Reductase Expression in Escherichia Coli written by Blake Alan Jacobson and published by . This book was released on 1997 with total page 308 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Methods for the Identification and Study of Required Genes in Escherichia Coli written by Roger Allyn Forsyth and published by . This book was released on 1999 with total page 300 pages. Available in PDF, EPUB and Kindle. Book excerpt: Required genes in Escherichia coli are those products which are essential for sustained growth and division. This dissertation describes the use of conditionally expressed antisense RNA to probe for genes required for growth under laboratory conditions. E. coli chromosomal fragments were cloned behind an IPTG inducible promoter. Out of 66,000 clones, 11 clones with inserts of 1 KB or less were sequenced, six of which produced RNA antisense to a portion of a chromosomally encoded mRNA. The antisense RNA's are complementary to mRNA's coding for ddlB (involved in peptidoglycan precursor synthesis), lepB (leader peptidase), and two previously unamed orfs. One orf, viaA (viability inhibited by antisense) was further characterized. ViaA is located in the periplasm and is a member of the thioredoxin family. Several interesting phenotypes associated with overexpression and loss of activity of ViaA are described. Another essential gene dnaA, required for initiating DNA replication, was studied using either a knockout with conditional plasmid complementation or inducible antisense. The first system verified that an excess of DnaA will increase the rate of chromosomal initiation events but that initiation events halt upon protein synthesis inhibition. DnaA overproduction also causes cell filamentation but not via an SOS mechanism. Regulation of dnaA was also studied using a dnaA-chb fusion integrated into the chromosome. A point mutation in the GUG start codon of dnaA to AUG revealed that the start codon utilization regulates the ultimate level of DnaA in the cell. Translation from both codons is correlated with growth rate although translation from the wildtype GUG start codon is lower than from the AUG mutated codon at all growth rates.

Download or read book Regulation of Several Genes Involved in DNA Replication in Escherichia Coli written by Thomas David Daws and published by . This book was released on 1988 with total page 272 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Studies on the Structure Function and Regulation of the DNA Replication Gene DnaA of Escherichia Coli K 12 written by Robert E. Braun and published by . This book was released on 1985 with total page 430 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book DNA Replication Control in Microbial Cell Factories written by Monika Glinkowska and published by Springer. This book was released on 2014-09-23 with total page 56 pages. Available in PDF, EPUB and Kindle. Book excerpt: This work describes the current knowledge of biochemical mechanisms regulating initiation of DNA replication in Escherichia coli, which focuses on the control of activity of the DnaA protein. Examples of direct linkages between DNA replication and other cellular processes are provided. In addition, similarities of the mechanisms of regulation of DNA replication operating in prokaryotic and eukaryotic cells are identified, and implications for understanding more complex processes, like carcinogenesis are suggested. Studies of recent years provided evidence that regulation of DNA replication in bacteria is more complex than previously anticipated. Multiple layers of control seem to ensure coordination of this process with the increase of cellular mass and the division cycle. Metabolic processes and membrane composition may serve as points where integration of genome replication with growth conditions occurs. It is also likely that coupling of DNA synthesis with cellular metabolism may involve interactions of replication proteins with other macromolecular complexes, responsible for various cellular processes. Thus, the exact set of factors participating in triggering the replication initiation may differ depending on growth conditions. Therefore, understanding the regulation of DNA duplication requires placing this process in the context of the current knowledge on bacterial metabolism, as well as cellular and chromosomal structure. Moreover, in both Escherichia coli and eukaryotic cells, replication initiator proteins were shown to play other roles in addition to driving the assembly of replication complexes, which constitutes another, yet not sufficiently understood, layer of coordinating DNA replication with the cell cycle.

Download or read book An Analysis of the Regulation of the ProS drpA Gene of Escherichia Coli written by Timothy T. Luby and published by . This book was released on 1991 with total page 244 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book FNR dependent Transcriptional Regulation in Escherichia Coli written by Eva Christine Ziegelhoffer and published by . This book was released on 1996 with total page 430 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book DNA Replication The Regulatory Mechanisms written by Patrick Hughes and published by Springer Science & Business Media. This book was released on 2012-12-06 with total page 429 pages. Available in PDF, EPUB and Kindle. Book excerpt: DNA replication is a key event in the cell cycle. Although our knowledge is far from complete and many elusive regulatory mechanisms still remain beyondour grasp, many enzymes and a multiplicity of biochemical mechanisms involved have been discovered. Recent findings in E. coli have confirmed and yet surpassed the original hypothesis of F. Jacob. In yeast and higher eucaryotes, the apparent redundancy in putative origins and initiators has made an estimation of the importance of each identified element difficult to access. In spite of well established methodologies - which are also described in the book - the origin identification in mammalian chromosomes is still a controversial subject. On the other hand, considerable advances have been made in our understanding of virus DNA replication and this continues to deepen and broaden our understanding of the controls of cellular DNA replication.

Download or read book Gene Regulation in Escherichia Coli Beyond the 0 rate0 Approximation written by Lok-hang So and published by . This book was released on 2011 with total page pages. Available in PDF, EPUB and Kindle. Book excerpt: The blueprint of a living cell is inscribed in its DNA. A region of DNA encoding a protein is called a gene. The cell reads the DNA and makes molecular machines made up of proteins to carry out all cellular functions required for survival. All cells live in ever-changing environments, and have different needs at different times. The control of when and how often each protein is produced from a gene is called gene regulation. Transcription, the copying of a DNA sequence into a complementary mRNA molecule, is the first step in the information flow from DNA to proteins, and most regulation is already done at the transcription level to avoid the production of superfluous intermediates. A living cell takes environmental stimuli as input, and regulates the activity of genes through DNA-binding proteins called transcription factors. The activity of a gene is described by its time-series of discrete mRNA production events. The events constituting this transcriptional time-series are stochastic and exhibit intermittent, bursty behavior, in bacteria as well as higher organisms. Thus the transcriptional time-series cannot be fully described by a simple chemical 0́−rate0́+0́4the probability per unit time of transcribing an mRNA molecule. An important consequence of this temporal complexity is that gene expression level can be tuned by varying different features of the time-series. It is then natural to ask: What modulation scheme is used by the cell to change expression levels of genes? Furthermore, if we look at the transcriptional time-series of multiple genes, would we see different modulation schemes for different genes, or a common modulation scheme shared by all genes? Last but not least, what is the molecular mechanism leading to bursty transcriptional time-series? What are the biophysical states that correspond to the active and inactive periods in a bursty transcriptional time-series? To answer these questions, I characterized the mRNA copy-number statistics from multiple promoters in the model organism Escherichia coli under various growth conditions using single-molecule fluorescence in situ hybridization. The kinetics of the underlying transcriptional time-series was then inferred using the two-state model, a simple stochastic mathematical model that describes bursty transcription time-series. I found that the degree of burstiness depends only on the gene expression level, while being independent of the details of gene regulation. The observed behavior is explained by the underlying variation in the duration of bursting events. At this stage, there is no mechanistic, molecular-level understanding of what gives rise to the bursty behavior of gene activity in bacteria. However, my finding here, that the properties of the transcriptional time-series are gene-independent rather than gene-specific, is contrary to the most common theoretical model used to explain bursty transcriptional time-series in bacteria, which involves the binding and unbinding of transcription factors at the promoter. My data suggests that the observed bursty kinetics arises from gene-nonspecific mechanisms such as DNA topology modulation, RNA polymerase dynamics, or regulation by broad-target DNA-binding proteins. Further investigation would narrow down the source of bursty transcriptional time-series.

Download or read book Gene Function written by Robert E. Glass and published by Univ of California Press. This book was released on 1982-01-01 with total page 516 pages. Available in PDF, EPUB and Kindle. Book excerpt: