Recombination Phenotypes of Escherichia coli greA Mutants

<p>Abstract</p> <p>Background</p> <p>The elongation factor GreA binds to RNA polymerase and modulates transcriptional pausing. Some recent research suggests that the primary role of GreA may not be to regulate gene expression, but rather, to promote the progression of replication forks which collide with RNA polymerase, and which might otherwise collapse. Replication fork collapse is known to generate dsDNA breaks, which can be recombinogenic. It follows that GreA malfunction could have consequences affecting homologous recombination.</p> <p>Results</p> <p><it>Escherichia coli </it>mutants bearing substitutions of the active site acidic residues of the transcription elongation factor GreA, D41N and E44K, were isolated as suppressors of growth inhibition by a toxic variant of the bacteriophage lambda Red-beta recombination protein. These mutants, as well as a D41A <it>greA </it>mutant and a <it>greA </it>deletion, were tested for proficiency in recombination events. The mutations were found to increase the efficiency of RecA-RecBCD-mediated and RecA-Red-mediated recombination, which are replication-independent, and to decrease the efficiency of replication-dependent Red-mediated recombination.</p> <p>Conclusion</p> <p>These observations provide new evidence for a role of GreA in resolving conflicts between replication and transcription.</p

Co-Orientation of Replication and Transcription Preserves Genome Integrity

In many bacteria, there is a genome-wide bias towards co-orientation of replication and transcription, with essential and/or highly-expressed genes further enriched co-directionally. We previously found that reversing this bias in the bacterium Bacillus subtilis slows replication elongation, and we proposed that this effect contributes to the evolutionary pressure selecting the transcription-replication co-orientation bias. This selection might have been based purely on selection for speedy replication; alternatively, the slowed replication might actually represent an average of individual replication-disruption events, each of which is counter-selected independently because genome integrity is selected. To differentiate these possibilities and define the precise forces driving this aspect of genome organization, we generated new strains with inversions either over ∼1/4 of the chromosome or at ribosomal RNA (rRNA) operons. Applying mathematical analysis to genomic microarray snapshots, we found that replication rates vary dramatically within the inverted genome. Replication is moderately impeded throughout the inverted region, which results in a small but significant competitive disadvantage in minimal medium. Importantly, replication is strongly obstructed at inverted rRNA loci in rich medium. This obstruction results in disruption of DNA replication, activation of DNA damage responses, loss of genome integrity, and cell death. Our results strongly suggest that preservation of genome integrity drives the evolution of co-orientation of replication and transcription, a conserved feature of genome organization

Electrophoretic deposition of graphene oxide on magnetic ribbon: Toward high sensitive and selectable magnetoimpedance response

Graphene oxide (GO) layers have shown to be fascinating elements for application in high performance sensors. They can be applied in multi-disciplinary designs based on surface selective sensing mechanisms. One immediate application of such surface sensitive elements is implementing of GO layer in magnetoimpedance (MI) sensors to improve their multi-functionality. In this paper, deposition of GO on the surface of Co-based amorphous ribbons (Co 68.15 Fe 4.35 Si 12.5 B 15 ) is performed using electrophoretic deposition (EPD) method to evaluate the MI response. MI ratio increased from 271% (bare ribbon) up to 281% and 301% EPD GO deposited within 4 and 8 min, respectively. Similar experiment for the ribbon drop coated with GO was carried out while no enhancement in MI response was seen. Vertical growth of GO on the surface of the ribbon in EPD and drop coated layers observed by topographical measurements. We explained the difference between the MI responses based on layers verticality and surface coverage. UV–Visible absorption and Raman spectroscopy were used to study the nature of GO. Gaining a high surface area of GO along with their biocompatible and anticorrosive properties atop the MI sensors can open pathways towards increasing applications of surface selective and high sensitive MI sensors

Electrophoretic deposition of graphene oxide on magnetic ribbon: Toward high sensitive and selectable magnetoimpedance response

Graphene oxide (GO) layers have shown to be fascinating elements for application in high performance sensors. They can be applied in multi-disciplinary designs based on surface selective sensing mechanisms. One immediate application of such surface sensitive elements is implementing of GO layer in magnetoimpedance (MI) sensors to improve their multi-functionality. In this paper, deposition of GO on the surface of Co-based amorphous ribbons (Co 68.15 Fe 4.35 Si 12.5 B 15 ) is performed using electrophoretic deposition (EPD) method to evaluate the MI response. MI ratio increased from 271% (bare ribbon) up to 281% and 301% EPD GO deposited within 4 and 8 min, respectively. Similar experiment for the ribbon drop coated with GO was carried out while no enhancement in MI response was seen. Vertical growth of GO on the surface of the ribbon in EPD and drop coated layers observed by topographical measurements. We explained the difference between the MI responses based on layers verticality and surface coverage. UV–Visible absorption and Raman spectroscopy were used to study the nature of GO. Gaining a high surface area of GO along with their biocompatible and anticorrosive properties atop the MI sensors can open pathways towards increasing applications of surface selective and high sensitive MI sensors

Electrophoretic deposition of graphene oxide on magnetic ribbon: Toward high sensitive and selectable magnetoimpedance response

Graphene oxide (GO) layers have shown to be fascinating elements for application in high performance sensors. They can be applied in multi-disciplinary designs based on surface selective sensing mechanisms. One immediate application of such surface sensitive elements is implementing of GO layer in magnetoimpedance (MI) sensors to improve their multi-functionality. In this paper, deposition of GO on the surface of Co-based amorphous ribbons (Co 68.15 Fe 4.35 Si 12.5 B 15 ) is performed using electrophoretic deposition (EPD) method to evaluate the MI response. MI ratio increased from 271% (bare ribbon) up to 281% and 301% EPD GO deposited within 4 and 8 min, respectively. Similar experiment for the ribbon drop coated with GO was carried out while no enhancement in MI response was seen. Vertical growth of GO on the surface of the ribbon in EPD and drop coated layers observed by topographical measurements. We explained the difference between the MI responses based on layers verticality and surface coverage. UV–Visible absorption and Raman spectroscopy were used to study the nature of GO. Gaining a high surface area of GO along with their biocompatible and anticorrosive properties atop the MI sensors can open pathways towards increasing applications of surface selective and high sensitive MI sensors

Effects on Growth by Changes of the Balance between GreA, GreB, and DksA Suggest Mutual Competition and Functional Redundancy in Escherichia coli

It is well known that ppGpp and DksA interact with bacterial RNA polymerase (RNAP) to alter promoter activity. This study suggests that GreA plays a major role and GreB plays a minor role in the ppGpp-DksA regulatory network. We present evidence that DksA and GreA/GreB are redundant and/or share similar functions: (i) on minimal medium GreA overproduction suppresses the growth defects of a dksA mutant; (ii) GreA and DksA overexpression partially suppresses the auxotrophy of a ppGpp-deficient strain; (iii) microarrays show that many genes are regulated similarly by GreA and DksA. We also find instances where GreA and DksA seem to act in opposition: (i) complete suppression of auxotrophy occurs by overexpression of GreA or DksA only in the absence of the other protein; (ii) PgadA and PgadE promoter fusions, along with many other genes, are dramatically affected in vivo by GreA overproduction only when DksA is absent; (iii) GreA and DksA show opposite regulation of a subset of genes. Mutations in key acidic residues of GreA and DksA suggest that properties seen here probably are not explained by known biochemical activities of these proteins. Our results indicate that the general pattern of gene expression and, in turn, the ability of Escherichia coli to grow under a defined condition are the result of a complex interplay between GreA, GreB, and DksA that also involves mutual control of their gene expression, competition for RNA polymerase binding, and similar or opposite action on RNA polymerase activity