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Multiple Ku orthologues mediate DNA non-homologous end-joining in the free-living form and during chronic infection of Sinorhizobium meliloti.

ABSTRACT: The bacterial non-homologous end-joining (NHEJ) apparatus is a two-component system that uses Ku and LigD to repair DNA double-strand breaks. Although the reaction mechanism has been extensively studied, much less is known about the physiological role of bacterial NHEJ. Recent studies suggest that NHEJ acts under conditions where DNA replication is reduced or absent (such as in a spore or stationary phase). Interestingly, genes encoding Ku and LigD have been identified in a wide range of bacteria that can chronically infect eukaryotic hosts. Strikingly, Sinohizobium meliloti, an intracellular symbiont of legume plants, carries four genes encoding Ku homologues (sku1 to sku4). Deletion analysis of the sku genes indicated that all Ku homologues are functional. One of these genes, sku2, is strongly expressed in free-living cells, as well as in bacteroid cells residing inside of the host plant. To visualize the NHEJ apparatus in vivo, SKu2 protein was fused to yellow fluorescent protein (YFP). Ionizing radiation (IR) induced focus formation of SKu2-YFP in free-living cells in a dosage-dependent manner. Moreover, SKu2-YFP foci formed in response to IR in non-dividing bacteroids, indicating that NHEJ system is functional even during the chronic infection phase of symbiosis.

SUBMITTER: Kobayashi H

PROVIDER: S-EPMC2770583 | biostudies-literature | 2008 Jan

REPOSITORIES: biostudies-literature

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Multiple Ku orthologues mediate DNA non-homologous end-joining in the free-living form and during chronic infection of Sinorhizobium meliloti.

Kobayashi Hajime H Simmons Lyle A LA Yuan Daniel S DS Broughton William J WJ Walker Graham C GC

Molecular microbiology 20071207 2

The bacterial non-homologous end-joining (NHEJ) apparatus is a two-component system that uses Ku and LigD to repair DNA double-strand breaks. Although the reaction mechanism has been extensively studied, much less is known about the physiological role of bacterial NHEJ. Recent studies suggest that NHEJ acts under conditions where DNA replication is reduced or absent (such as in a spore or stationary phase). Interestingly, genes encoding Ku and LigD have been identified in a wide range of bacteri ...[more]

PMID: 18067541

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Project description:The proposal aims to characterise the pathways of DSB repair and recombination in Arabidopsis with the main focus of this research being the NHEJ pathway of illegitimate recombination. We will build on our expertise and resources in the field of DSB repair in plants, using the Arabidopsis NHEJ mutant atku80 which is an excellent model system for the study of DSB repair in higher eukaryotes (West et al., 2002 Plant J. 31, 517-28). Comparison of the transcriptome in NHEJ mutant and wild type plants under different experimental conditions will identify novel candidate genes involved in DNA DSB repair or damage signalling pathways.We will conduct 4 separate experiments on Arabidopsis seedlings grown on 0.5 MS media: the first will be a control consisting of Wassilewskija (WS-2) plants grown under standard conditions. In the second experiment WS plants will be exposed to the chemotherapeutic agent bleomycin (1 microgram per ml). This agent has been shown to cause both single and double strand breaks in the genome of Arabidopsis seedlings (Menke et al., 2001 Mut. Res. 493, 87-93). This agent is used in preference to gamma irradiation, which causes high levels of oxidative damage to cellular components. These experiments will characterise for the first time the transcriptional responses of Arabidopsis cells to the specific induction of DNA strand breaks. The transcript profile will also be determined in untreated atku80 mutants. This experiment will identify the components of novel and previously characterised DNA repair pathways up regulated in the mutant background. Finally, transcript analysis of bleomycin treated atku80 mutants and comparison with untreated mutant plants and both untreated and treated WS plants will identify novel putative DSB repair genes up regulated in response to genotoxic stress in the absence of a Ku80 mediated DSB repair pathway.The results of these studies will provide new and important information which will be instrumental in identifying novel genes and pathways involved in DNA repair and genome stability in plants and help elucidate the fundamental differences that exist between animal and plant DSB repair processes. Experiment Overall Design: Number of plants pooled:20

Dataset Information

Multiple Ku orthologues mediate DNA non-homologous end-joining in the free-living form and during chronic infection of Sinorhizobium meliloti.

Publications

Multiple Ku orthologues mediate DNA non-homologous end-joining in the free-living form and during chronic infection of Sinorhizobium meliloti.

Similar Datasets

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