Transcriptional termination at a fully rho-independent site in Escherichia coli is prevented by uninterrupted translation of the nascent RNA.

1987 ◽  
Vol 6 (4) ◽  
pp. 1115-1119 ◽  
Author(s):  
J.J. Wright ◽  
R.S. Hayward
Keyword(s):  
Escherichia Coli ◽  
Transcriptional Termination ◽  
Nascent Rna ◽  
Independent Site

scholarly journals POLARITY IN SEGMENTS OF THE ESCHERICHIA COLI trp OPERON WITH DELETED INTRAOPERONIC TRANSLATIONAL INITIATION SIGNALS

Genetics ◽  
1975 ◽  
Vol 80 (4) ◽  
pp. 651-666
Author(s):  
Yasunobu Kano ◽  
Fumio Imamoto
Keyword(s):  
Escherichia Coli ◽  
Translation Initiation ◽  
Nonsense Mutation ◽  
Messenger Rna ◽  
Distal Part ◽  
High Efficiency ◽  
Translational Initiation ◽  
Transcriptional Termination ◽  
Trp Operon

ABSTRACT The effect of deletion of the operator-distal genes of the trp operon, including the trpE-trpD intercistronic punctuation point, on the degree of transcriptional polarity (in this case the effect of a nonsense mutation on the level of mRNA from the distal part of the very gene where the mutation is located) was investigated. Double mutants which contain a nonsense mutation and a deletion in trpE were constructed, and the degree of transcriptional polarity was estimated by the decrease in messenger RNA for the operator-distal trpE beyond the nonsense mutation, as well as by the production of truncated messenger RNA for the region of trpE proximal to the nonsense mutation. The content of mRNA of operator-distal trpE and the size of the mRNA of operator-proximal trpE of the double mutants show that transcriptional polarity is not relaxed as a function of distance of the nonsense mutation from the operator-distal end of the trpE segment (at which the subsequent high efficiency translational initiation signal has been deleted). These findings are consistent with the conclusion that the degree of polarity depends on the distance of the nonsense mutation fro mthe subsequent translation initiation signal, but not on its distance from the operator-distal end, including possible translational or transcriptional termination signals

scholarly journals The Role of Replication Clamp-Loader Protein HolC of Escherichia coli in Overcoming Replication/Transcription Conflicts

mBio ◽  
2021 ◽  
Vol 12 (2) ◽  
Author(s):  
Deani L. Cooper ◽  
Taku Harada ◽  
Samia Tamazi ◽  
Alexander E. Ferrazzoli ◽  
Susan T. Lovett
Keyword(s):  
Escherichia Coli ◽  
Dna Replication ◽  
Rna Polymerase ◽  
Transcription Elongation ◽  
Clamp Loader ◽  
Content Type ◽  
Growth Defects ◽  
Transcriptional Termination ◽  
Transcription Complexes

ABSTRACT In Escherichia coli, DNA replication is catalyzed by an assembly of proteins, the DNA polymerase III holoenzyme. This complex includes the polymerase and proofreading subunits, the processivity clamp, and clamp loader complex. The holC gene encodes an accessory protein (known as χ) to the core clamp loader complex and is the only protein of the holoenzyme that binds to single-strand DNA binding protein, SSB. HolC is not essential for viability, although mutants show growth impairment, genetic instability, and sensitivity to DNA damaging agents. In this study, we isolate spontaneous suppressor mutants in a ΔholC strain and identify these by whole-genome sequencing. Some suppressors are alleles of RNA polymerase, suggesting that transcription is problematic for holC mutant strains, or alleles of sspA, encoding stringent starvation protein. Using a conditional holC plasmid, we examine factors affecting transcription elongation and termination for synergistic or suppressive effects on holC mutant phenotypes. Alleles of RpoA (α), RpoB (β), and RpoC (β′) RNA polymerase holoenzyme can partially suppress loss of HolC. In contrast, mutations in transcription factors DksA and NusA enhanced the inviability of holC mutants. HolC mutants showed enhanced sensitivity to bicyclomycin, a specific inhibitor of Rho-dependent termination. Bicyclomycin also reverses suppression of holC by rpoA, rpoC, and sspA. An inversion of the highly expressed rrnA operon exacerbates the growth defects of holC mutants. We propose that transcription complexes block replication in holC mutants and that Rho-dependent transcriptional termination and DksA function are particularly important to sustain viability and chromosome integrity. IMPORTANCE Transcription elongation complexes present an impediment to DNA replication. We provide evidence that one component of the replication clamp loader complex, HolC, of Escherichia coli is required to overcome these blocks. This genetic study of transcription factor effects on holC growth defects implicates Rho-dependent transcriptional termination and DksA function as critical. It also implicates, for the first time, a role of SspA, stringent starvation protein, in avoidance or tolerance of replication/replication conflicts. We speculate that HolC helps avoid or resolve collisions between replication and transcription complexes, which become toxic in HolC’s absence.

scholarly journals The role of replication clamp-loader protein HolC of Escherichia coli in overcoming replication / transcription conflicts

2020 ◽  
Author(s):  
Deani L. Cooper ◽  
Taku Harada ◽  
Samia Tamazi ◽  
Alexander E. Ferrazzoli ◽  
Susan T. Lovett
Keyword(s):  
Escherichia Coli ◽  
Dna Replication ◽  
Rna Polymerase ◽  
Transcription Elongation ◽  
Specific Inhibitor ◽  
Clamp Loader ◽  
Enhanced Sensitivity ◽  
Transcriptional Termination ◽  
Transcription Complexes

ABSTRACTIn Escherichia coli, DNA replication is catalyzed by an assembly of proteins, the DNA polymerase III holoenzyme. This complex includes the polymerase and proofreading subunits as well as the processivity clamp and clamp loader complex. The holC gene encodes an accessory protein (known as x) to the core clamp loader complex and is the only protein of the holoenzyme that binds to single-strand DNA binding protein, SSB. HolC is not essential for viability although mutants show growth impairment, genetic instability and sensitivity to DNA damaging agents. In this study, to elucidate the role of HolC in replication, we isolate spontaneous suppressor mutants in a holCΔ strain and identify these by whole genome sequencing. Some suppressors are alleles of RNA polymerase, suggesting that transcription is problematic for holC mutant strains or sspA, stringent starvation protein. Using a conditional holC plasmid, we examine factors affecting transcription elongation and termination for synergistic or suppressive effects on holC mutant phenotypes. Alleles of RpoA (α), RpoB (β) and RpoC (β’) RNA polymerase holoenzyme can partially suppress loss of HolC. In contrast, mutations in transcription factors DksA and NusA enhanced the inviability of holC mutants. Mfd had no effect nor did elongation factors GreA and GreB. HolC mutants showed enhanced sensitivity to bicyclomycin, a specific inhibitor of Rho-dependent termination. Bicyclomycin also reverses suppression of holC by rpoA rpoC and sspA.These results are consistent with the hypothesis that transcription complexes block replication in holC mutants and Rho-dependent transcriptional termination and DksA function are particularly important to sustain viability and chromosome integrity.IMPORTANCETranscription elongation complexes present an impediment to DNA replication. We provide evidence that one component of the replication clamp loader complex, HolC, of E. coli is required to overcome these blocks. This genetic study of transcription factor effects on holC growth defects implicates Rho-dependent transcriptional termination and DksA function as critical. It also implicates, for the first time, a role of SspA, stringent starvation protein, in avoidance or tolerance of replication/replication conflicts. We speculate that HolC helps resolve codirectional collisions between replication and transcription complexes, which become toxic in HolC’s absence.

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