Affiliation:
1. Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina, USA
Abstract
ABSTRACT
dGTP starvation, a newly discovered phenomenon in which
Escherichia coli
cells are starved specifically for the DNA precursor dGTP, leads to impaired growth and, ultimately, cell death. Phenomenologically, it represents an example of nutritionally induced unbalanced growth: cell mass amplifies normally as dictated by the nutritional status of the medium, but DNA content growth is specifically impaired. The other known example of such a condition, thymineless death (TLD), involves starvation for the DNA precursor dTTP, which has been found to have important chemotherapeutic applications. Experimentally, dGTP starvation is induced by depriving an
E. coli
gpt optA1
strain of its required purine source, hypoxanthine. In our studies of this phenomenon, we noted the emergence of a relatively high frequency of suppressor mutants that proved resistant to the treatment. To study such suppressors, we used next-generation sequencing on a collection of independently obtained mutants. A significant fraction was found to carry a defect in the PurR transcriptional repressor, controlling
de novo
purine biosynthesis, or in its downstream
purEK
operon. Thus, upregulation of
de novo
purine biosynthesis appears to be a major mode of overcoming the lethal effects of dGTP starvation. In addition, another large fraction of the suppressors contained a large tandem duplication of a 250- to 300-kb genomic region that included the
purEK
operon as well as the
acrAB
-encoded multidrug efflux system. Thus, the suppressive effects of the duplications could potentially involve beneficial effects of a number of genes/operons within the amplified regions.
IMPORTANCE
Concentrations of the four precursors for DNA synthesis (2′-deoxynucleoside-5′-triphosphates [dNTPs]) are critical for both the speed of DNA replication and its accuracy. Previously, we investigated consequences of dGTP starvation, where the DNA precursor dGTP was specifically reduced to a low level. Under this condition,
E. coli
cells continued cell growth but eventually developed a DNA replication defect, leading to cell death due to formation of unresolvable DNA structures. Nevertheless, dGTP-starved cultures eventually resumed growth due to the appearance of resistant mutants. Here, we used whole-genome DNA sequencing to identify the responsible suppressor mutations. We show that the majority of suppressors can circumvent death by upregulating purine
de novo
biosynthesis, leading to restoration of dGTP to acceptable levels.
Funder
HHS | NIH | National Institute of Environmental Health Sciences
Publisher
American Society for Microbiology
Subject
Molecular Biology,Microbiology
Reference61 articles.
1. Maaløe O Kjeldgaard NO . 1966. Control of macromolecular synthesis: a study of DNA, RNA, and protein synthesis, in bacteria. W. A. Benjamin, New York, NY.
2. Dependency on Medium and Temperature of Cell Size and Chemical Composition during Balanced Growth of Salmonella typhimurium
3. Helmstetter CE . 1996. Timing of synthetic activities in the cell cycle, p 1627–1639. InNeidhardtFCCurtissR IIIIngrahamJLLinECCLowKBMagasanikBReznikoffWSRileyMSchaechterMUmbargerH (ed), Escherichia coli and Salmonella: cellular and molecular biology, 2nd ed.American Society for Microbiology, Washington, DC.
4. On the Bacterial Life Sequence
5. Regulating DNA Replication in Bacteria
Cited by
1 articles.
订阅此论文施引文献
订阅此论文施引文献,注册后可以免费订阅5篇论文的施引文献,订阅后可以查看论文全部施引文献