Resolution of head-on collisions between the transcription machinery and bacteriophage Phi 29 DNA polymerase is dependent on RNA polymerase translocation

Elías‐Arnanz, Montserrat; Salas, Margarita

doi:10.1093/emboj/18.20.5675

Cited by 33 publications

(37 citation statements)

References 38 publications

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“…Since the E. coli chromosome seems to prefer the codirectional arrangement (2), it was proposed that head-on collisions are more detrimental for the chromosomal replication than codirectional ones (3). While some in vitro and in vivo experiments supported this idea (14,25,26,39), other studies observed replication stalling for both types of collisions (12,13) or neither of them (34). Here, we revisited this matter by using electrophoresis analysis of replication intermediates for ColE1-type plasmids in vivo.…”

Section: Discussionmentioning

confidence: 89%

“…When RNA polymerase was allowed to move slowly, replication fork pausing became less pronounced. Phage 29 replisome, in contrast, halted upon encountering stalled B. subtilis RNA polymerase in both orientations (12,13). Once RNA polymerase movement was resumed, DNA synthesis continued with its normal speed in the head-on case but was much slower for the codirectional alignment.…”

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confidence: 99%

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Mechanisms of Transcription-Replication Collisions in Bacteria

Mirkin

2005

Molecular and Cellular Biology

162

153

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While collisions between replication and transcription in bacteria are deemed inevitable, the fine details of the interplay between the two machineries are poorly understood. In this study, we evaluate the effects of transcription on the replication fork progression in vivo, by using electrophoresis analysis of replication intermediates. Studying Escherichia coli plasmids, which carry constitutive or inducible promoters in different orientations relative to the replication origin, we show that the mutual orientation of the two processes determines their mode of interaction. Replication elongation appears not to be affected by transcription proceeding in the codirectional orientation. Head-on transcription, by contrast, leads to severe inhibition of the replication fork progression. Furthermore, we evaluate the mechanism of this inhibition by limiting the area of direct contact between the two machineries. We observe that replication pausing zones coincide exactly with transcribed DNA segments. We conclude, therefore, that the replication fork is most likely attenuated upon direct physical interaction with the head-on transcription machinery.In bacteria, DNA replication and transcription continue throughout the life cycle. The speed of the replication fork progression in Escherichia coli is ϳ1,000 bp per s (16), while the elongation rate for RNA polymerase is just 50 nucleotides (nt) per s (15); i.e., replication is approximately 20-fold faster than transcription. Since the two processes proceed simultaneously, frequent collisions between replication and transcription seem unavoidable (3, 38). Given that both processes are polar, the collisions can occur either head-on or codirectionally (Fig. 1). In the case of head-on collisions, the front edge of RNA polymerase meets the hexameric DNA helicase DnaB that moves along the lagging strand template. In the codirectional case, by contrast, the front edge of the leading strand DNA polymerase collides with the rear edge of RNA polymerase.While it is unclear a priori which type of collision is more damaging for DNA metabolism, the data on the organization of bacterial genomes point to selection against head-on collisions. Sequencing of the E. coli genome revealed that there is a bias towards codirectional alignment of transcription units with replication (2). Most strikingly, all seven ribosomal operons face the direction of their replichores. For other genes, however, this bias is much less pronounced: ϳ62% of tRNA genes and ϳ55% of protein-coding genes are aligned codirectionally with replication. Similar principles of gene arrangement were observed for other bacteria, such as Bacillus subtilis, Borrelia burgdorferi, Treponema pallidum, Haemophilus influenzae, Helicobacter pylori, Mycoplasma genitalium, and Mycoplasma pneumoniae (36), as well as for bacteriophages T7 and lambda (3).Experimental data on transcription-replication collision are relatively scarce. This problem was addressed in vitro by studying interactions between bacterial RNA polymerases and phage repl...

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Section: Discussionmentioning

confidence: 89%

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confidence: 99%

Mechanisms of Transcription-Replication Collisions in Bacteria

Mirkin

2005

Molecular and Cellular Biology

162

153

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“…DNA polymerase from 29 can bypass B. subtilis RNAP during head-on encounters. During codirectional encounters, the 29 DNA polymerase slows down and follows RNAP (35,36).…”

Section: Discussionmentioning

confidence: 99%

Genome-wide coorientation of replication and transcription reduces adverse effects on replication in Bacillus subtilis

Wang

Berkmen

Grossman

2007

Proc. Natl. Acad. Sci. U.S.A.

102

119

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In many bacteria, there is a strong bias for genes to be encoded on the leading strand of DNA, resulting in coorientation of replication and transcription. In Bacillus subtilis, transcription of the majority of genes (75%) is cooriented with replication. By using genomewide profiling of replication with DNA microarrays, we found that this coorientation bias reduces adverse effects of transcription on replication. We found that in wild-type cells, transcription did not appear to affect the rate of replication elongation. However, in mutants with reversed transcription bias for an extended region of the chromosome, replication elongation was slower. This reduced replication rate depended on transcription and was limited to the region in which the directions of replication and transcription are opposed. These results support the hypothesis that the strong bias to coorient transcription and replication is due to selective pressure for processive, efficient, and accurate replication.DNA microarrays ͉ elongation of replication ͉ genomic organization ͉ genomic stability ͉ origin of replication M any aspects of the organization of bacterial genomes are conserved and important for cell survival. DNA rearrangements, including large chromosomal inversions, can lead to inviability, decreased fitness, and impaired development (1-4). It has been proposed that genomic organization affects replication, transcription, and segregation of genomes (5).One benefit of proper genomic organization may be the reduction of potential conflicts between replication and transcription (5-7). The same DNA template is used by RNA polymerase (RNAP) for transcription and by the replisome (DNA polymerase and associated proteins) for replication. Transcription complexes that are stalled, initiating, or terminating can slow or block replication (8, 9). RNAP and the replisome can collide when moving toward each other (head-on), or when the replisome, which moves faster than RNAP, catches up with RNAP moving in the same direction (codirectional). Head-on and codirectional collisions can occur when genes are on the lagging and leading strands, respectively.The consequences of head-on and codirectional collisions are different. In Escherichia coli, high levels of transcription can effectively slow or block progression of replication forks if transcription and replication are head-on, but not if they are codirectional (10). In French's landmark study, an inducible plasmid-derived origin of replication was positioned on either side of an rRNA operon, one of the most highly transcribed operons. Replication fork progression, monitored by EM, was much slower when running against, rather than with, the direction of transcription. Plasmid DNA replication in E. coli can also be hindered by head-on transcription from a strong inducible promoter, probably because of collisions between the replisome and RNAP (ref.

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“…In this phage cooriented collisions lead, as in E. coli, to replication slowdown, which is probably caused by the DNAP following an actively transcribing RNAP (Elias-Arnanz & Salas, 1997). On the other hand, head-on collisions are dealt with in such a way that the DNAP by-passes RNAP without displacing it, in a mechanism that is probably different from that of T4 (Elias-Arnanz & Salas, 1999). It remains to be investigated whether bacteria lacking replication bias have also found ways of dealing with the problem of collision between polymerases.…”

Section: Understanding Replication From the Organization It Inducesmentioning

confidence: 99%

The replication-related organization of bacterial genomes

2004

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The replication of the chromosome is among the most essential functions of the bacterial cell and influences many other cellular mechanisms, from gene expression to cell division. Yet the way it impacts on the bacterial chromosome was not fully acknowledged until the availability of complete genomes allowed one to look upon genomes as more than bags of genes. Chromosomal replication includes a set of asymmetric mechanisms, among which are a division in a lagging and a leading strand and a gradient between early and late replicating regions. These differences are the causes of many of the organizational features observed in bacterial genomes, in terms of both gene distribution and sequence composition along the chromosome. When asymmetries or gradients increase in some genomes, e.g. due to a different composition of the DNA polymerase or to a higher growth rate, so do the corresponding biases. As some of the features of the chromosome structure seem to be under strong selection, understanding such biases is important for the understanding of chromosome organization and adaptation. Inversely, understanding chromosome organization may shed further light on questions relating to replication and cell division. Ultimately, the understanding of the interplay between these different elements will allow a better understanding of bacterial genetics and evolution.

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Resolution of head-on collisions between the transcription machinery and bacteriophage Phi 29 DNA polymerase is dependent on RNA polymerase translocation

Cited by 33 publications

References 38 publications

Mechanisms of Transcription-Replication Collisions in Bacteria

Mechanisms of Transcription-Replication Collisions in Bacteria

Genome-wide coorientation of replication and transcription reduces adverse effects on replication in Bacillus subtilis

The replication-related organization of bacterial genomes

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