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Dynamics and Cell-Type Specificity of the DNA Double-Strand Break Repair Protein RecN in the Developmental Cyanobacterium Anabaena sp. Strain PCC 7120.

Hu S, Wang J, Wang L, Zhang CC, Chen WL - PLoS ONE (2015)

Bottom Line: Treatment by a drug causing DNA double-strand breaks (DSBs) induced reorganization of the RecN focus preferentially towards the mid-cell position.RecN-GFP was absent in most mature heterocysts.Furthermore, our results showed that HetR, a central player in heterocyst development, was involved in the proper positioning and distribution of RecN-GFP.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, 430070 Wuhan, China.

ABSTRACT
DNA replication and repair are two fundamental processes required in life proliferation and cellular defense and some common proteins are involved in both processes. The filamentous cyanobacterium Anabaena sp. strain PCC 7120 is capable of forming heterocysts for N2 fixation in the absence of a combined-nitrogen source. This developmental process is intimately linked to cell cycle control. In this study, we investigated the localization of the DNA double-strand break repair protein RecN during key cellular events, such as chromosome damaging, cell division, and heterocyst differentiation. Treatment by a drug causing DNA double-strand breaks (DSBs) induced reorganization of the RecN focus preferentially towards the mid-cell position. RecN-GFP was absent in most mature heterocysts. Furthermore, our results showed that HetR, a central player in heterocyst development, was involved in the proper positioning and distribution of RecN-GFP. These results showed the dynamics of RecN in DSB repair and suggested a differential regulation of DNA DSB repair in vegetative cell and heterocysts. The absence of RecN in mature heterocysts is compatible with the terminal nature of these cells.

No MeSH data available.


Related in: MedlinePlus

RecN localization is affected in patS or hetR mutant.(A) RecN-GFP foci in the strain RG-PM cultured in the medium BG11. (B) RecN-GFP foci in the strain RG-PM cultured in the medium BG110. The red arrows indicate heterocysts. (C) The localization of RecN-GFP foci in the strain RG-PM cultured in different media. The coordinate 0 is the center of the cell. The statistical method used here was the same with that in Fig 1B. (D) RecN-GFP foci in the strain RG-HM cultured in the medium BG11. (E) RecN-GFP foci in RG-HM cultured in the medium BG110, the red arrow indicates a cell with 2 foci. (F) The localization of RecN-GFP foci in the strain RG-HM cultured in different media. The coordinate 0 is the center of the cell. The statistical method used here was the same with that in Fig 1B. Photographs were taken by using an Olympus FV1000 confocal microscope. Cells were stained with DAPI (blue). The red fluorescence is from the photosynthetic pigments. Scale bars correspond to 1 μm.
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pone.0139362.g006: RecN localization is affected in patS or hetR mutant.(A) RecN-GFP foci in the strain RG-PM cultured in the medium BG11. (B) RecN-GFP foci in the strain RG-PM cultured in the medium BG110. The red arrows indicate heterocysts. (C) The localization of RecN-GFP foci in the strain RG-PM cultured in different media. The coordinate 0 is the center of the cell. The statistical method used here was the same with that in Fig 1B. (D) RecN-GFP foci in the strain RG-HM cultured in the medium BG11. (E) RecN-GFP foci in RG-HM cultured in the medium BG110, the red arrow indicates a cell with 2 foci. (F) The localization of RecN-GFP foci in the strain RG-HM cultured in different media. The coordinate 0 is the center of the cell. The statistical method used here was the same with that in Fig 1B. Photographs were taken by using an Olympus FV1000 confocal microscope. Cells were stained with DAPI (blue). The red fluorescence is from the photosynthetic pigments. Scale bars correspond to 1 μm.

Mentions: Our data showed that a RecN focus was present at the early stage of heterocyst differentiation, but disappeared in most mature heterocysts. We further performed RecN-GFP localization in a patS mutant UHM114 [39], where the patS gene was deleted, and a hetR mutant hetR216 carrying a loss-of-function point mutation [40,41]. The patS gene encodes an inhibitor of heterocyst differentiation, while hetR encodes a transcription factor required for heterocyst development [24,25]. Two genes, patS and hetR, are involved in heterocyst differentiation and patterning in Anabaena [42,43]. Under nitrogen deprivation conditions, UHM114 develops into heterocysts at a higher frequency as compared to the wild type strain of Anabaena, whereas hetR216 has no capability to form heterocysts [25]. The UHM114 strain with expressed RecN-GFP is named as RG-PM, while the hetR216 strain with expressed RecN-GFP is named as RG-HM. When cultured in BG11 medium containing a combined nitrogen, both RG-PM and RG-HM showed a similar location of RecN-GFP foci to that observed in RG-W (wild-type contrast), displaying a single discrete focus at DNA-free parts of the cells (Fig 6A and 6D). In the nitrogen–deprivation medium BG110, RG-PM developed more heterocysts as expected, but the localization of RecN-GFP foci was not different from that in RG-W (Fig 6B and 6C). However, the RG-HM strain showed two differences from RG-W under the same culture conditions. First, RecN-GFP foci in RG-HM cells were mostly colocalized with nucleoid-occupied regions at the center area of cells (Fig 6E and 6F). Secondly, 12.3% of the cells had 2 or 3 foci (Fig 6E). The phenomenon was observed in RG-W cells only after treatment by Mitomycin C.


Dynamics and Cell-Type Specificity of the DNA Double-Strand Break Repair Protein RecN in the Developmental Cyanobacterium Anabaena sp. Strain PCC 7120.

Hu S, Wang J, Wang L, Zhang CC, Chen WL - PLoS ONE (2015)

RecN localization is affected in patS or hetR mutant.(A) RecN-GFP foci in the strain RG-PM cultured in the medium BG11. (B) RecN-GFP foci in the strain RG-PM cultured in the medium BG110. The red arrows indicate heterocysts. (C) The localization of RecN-GFP foci in the strain RG-PM cultured in different media. The coordinate 0 is the center of the cell. The statistical method used here was the same with that in Fig 1B. (D) RecN-GFP foci in the strain RG-HM cultured in the medium BG11. (E) RecN-GFP foci in RG-HM cultured in the medium BG110, the red arrow indicates a cell with 2 foci. (F) The localization of RecN-GFP foci in the strain RG-HM cultured in different media. The coordinate 0 is the center of the cell. The statistical method used here was the same with that in Fig 1B. Photographs were taken by using an Olympus FV1000 confocal microscope. Cells were stained with DAPI (blue). The red fluorescence is from the photosynthetic pigments. Scale bars correspond to 1 μm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4592062&req=5

pone.0139362.g006: RecN localization is affected in patS or hetR mutant.(A) RecN-GFP foci in the strain RG-PM cultured in the medium BG11. (B) RecN-GFP foci in the strain RG-PM cultured in the medium BG110. The red arrows indicate heterocysts. (C) The localization of RecN-GFP foci in the strain RG-PM cultured in different media. The coordinate 0 is the center of the cell. The statistical method used here was the same with that in Fig 1B. (D) RecN-GFP foci in the strain RG-HM cultured in the medium BG11. (E) RecN-GFP foci in RG-HM cultured in the medium BG110, the red arrow indicates a cell with 2 foci. (F) The localization of RecN-GFP foci in the strain RG-HM cultured in different media. The coordinate 0 is the center of the cell. The statistical method used here was the same with that in Fig 1B. Photographs were taken by using an Olympus FV1000 confocal microscope. Cells were stained with DAPI (blue). The red fluorescence is from the photosynthetic pigments. Scale bars correspond to 1 μm.
Mentions: Our data showed that a RecN focus was present at the early stage of heterocyst differentiation, but disappeared in most mature heterocysts. We further performed RecN-GFP localization in a patS mutant UHM114 [39], where the patS gene was deleted, and a hetR mutant hetR216 carrying a loss-of-function point mutation [40,41]. The patS gene encodes an inhibitor of heterocyst differentiation, while hetR encodes a transcription factor required for heterocyst development [24,25]. Two genes, patS and hetR, are involved in heterocyst differentiation and patterning in Anabaena [42,43]. Under nitrogen deprivation conditions, UHM114 develops into heterocysts at a higher frequency as compared to the wild type strain of Anabaena, whereas hetR216 has no capability to form heterocysts [25]. The UHM114 strain with expressed RecN-GFP is named as RG-PM, while the hetR216 strain with expressed RecN-GFP is named as RG-HM. When cultured in BG11 medium containing a combined nitrogen, both RG-PM and RG-HM showed a similar location of RecN-GFP foci to that observed in RG-W (wild-type contrast), displaying a single discrete focus at DNA-free parts of the cells (Fig 6A and 6D). In the nitrogen–deprivation medium BG110, RG-PM developed more heterocysts as expected, but the localization of RecN-GFP foci was not different from that in RG-W (Fig 6B and 6C). However, the RG-HM strain showed two differences from RG-W under the same culture conditions. First, RecN-GFP foci in RG-HM cells were mostly colocalized with nucleoid-occupied regions at the center area of cells (Fig 6E and 6F). Secondly, 12.3% of the cells had 2 or 3 foci (Fig 6E). The phenomenon was observed in RG-W cells only after treatment by Mitomycin C.

Bottom Line: Treatment by a drug causing DNA double-strand breaks (DSBs) induced reorganization of the RecN focus preferentially towards the mid-cell position.RecN-GFP was absent in most mature heterocysts.Furthermore, our results showed that HetR, a central player in heterocyst development, was involved in the proper positioning and distribution of RecN-GFP.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, 430070 Wuhan, China.

ABSTRACT
DNA replication and repair are two fundamental processes required in life proliferation and cellular defense and some common proteins are involved in both processes. The filamentous cyanobacterium Anabaena sp. strain PCC 7120 is capable of forming heterocysts for N2 fixation in the absence of a combined-nitrogen source. This developmental process is intimately linked to cell cycle control. In this study, we investigated the localization of the DNA double-strand break repair protein RecN during key cellular events, such as chromosome damaging, cell division, and heterocyst differentiation. Treatment by a drug causing DNA double-strand breaks (DSBs) induced reorganization of the RecN focus preferentially towards the mid-cell position. RecN-GFP was absent in most mature heterocysts. Furthermore, our results showed that HetR, a central player in heterocyst development, was involved in the proper positioning and distribution of RecN-GFP. These results showed the dynamics of RecN in DSB repair and suggested a differential regulation of DNA DSB repair in vegetative cell and heterocysts. The absence of RecN in mature heterocysts is compatible with the terminal nature of these cells.

No MeSH data available.


Related in: MedlinePlus