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Regulation and role of Arabidopsis CUL4-DDB1A-DDB2 in maintaining genome integrity upon UV stress.

Molinier J, Lechner E, Dumbliauskas E, Genschik P - PLoS Genet. (2008)

Bottom Line: Moreover, we provide evidences for crosstalks between GGR, the plant-specific photo reactivation pathway and the RAD1-RAD10 endonucleases upon UV exposure.Finally, we report that DDB2 degradation upon UV stress depends not only on CUL4, but also on the checkpoint protein kinase Ataxia telangiectasia and Rad3-related (ATR).Interestingly, we found that DDB1A shuttles from the cytoplasm to the nucleus in an ATR-dependent manner, highlighting an upstream level of control and a novel mechanism of regulation of this E3 ligase.

View Article: PubMed Central - PubMed

Affiliation: Institut de Biologie Moléculaire des Plantes du CNRS (UPR2357), conventionné avec l'Université Louis Pasteur, Strasbourg, France.

ABSTRACT
Plants use the energy in sunlight for photosynthesis, but as a consequence are exposed to the toxic effect of UV radiation especially on DNA. The UV-induced lesions on DNA affect both transcription and replication and can also have mutagenic consequences. Here we investigated the regulation and the function of the recently described CUL4-DDB1-DDB2 E3 ligase in the maintenance of genome integrity upon UV-stress using the model plant Arabidopsis. Physiological, biochemical, and genetic evidences indicate that this protein complex is involved in global genome repair (GGR) of UV-induced DNA lesions. Moreover, we provide evidences for crosstalks between GGR, the plant-specific photo reactivation pathway and the RAD1-RAD10 endonucleases upon UV exposure. Finally, we report that DDB2 degradation upon UV stress depends not only on CUL4, but also on the checkpoint protein kinase Ataxia telangiectasia and Rad3-related (ATR). Interestingly, we found that DDB1A shuttles from the cytoplasm to the nucleus in an ATR-dependent manner, highlighting an upstream level of control and a novel mechanism of regulation of this E3 ligase.

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Genetic interactions in NER and HR.One-week-old single and double mutant plants (A) ddb1a-2-ddb2-2; cul4-1-cen2-2; ddb1a-2-cen2-2; ddb2-2-cen2-2 (B) cul4-1-rad10; ddb1a-2-rad10; ddb2-2-rad10; cen2-2-rad1 were exposed to 600 J/m2 of UV-C. Root growth was measured 24h following irradiation. Root growth was calculated relative to the corresponding untreated plants (±SEM). Eight plants per replicate were used and experiments were triplicated. For all the single and double mutants p<0.05, compared to WT plants. Because ddb2-2 is in a different Arabidopsis ecotype, the single control mutants were selected as segregants from each double mutant involving ddb2-2 and are indicated by *.
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pgen-1000093-g003: Genetic interactions in NER and HR.One-week-old single and double mutant plants (A) ddb1a-2-ddb2-2; cul4-1-cen2-2; ddb1a-2-cen2-2; ddb2-2-cen2-2 (B) cul4-1-rad10; ddb1a-2-rad10; ddb2-2-rad10; cen2-2-rad1 were exposed to 600 J/m2 of UV-C. Root growth was measured 24h following irradiation. Root growth was calculated relative to the corresponding untreated plants (±SEM). Eight plants per replicate were used and experiments were triplicated. For all the single and double mutants p<0.05, compared to WT plants. Because ddb2-2 is in a different Arabidopsis ecotype, the single control mutants were selected as segregants from each double mutant involving ddb2-2 and are indicated by *.

Mentions: CENTRIN2 (CEN2) and the heterodimer RAD1-RAD10 (XPF-ERCC1) act in the GGR-NER repair pathway [7]. CEN2 is part of the XPC-RAD23 recognition complex, whereas, RAD1-RAD10 in cooperation with RAD2 are the endonucleases excising the bulky DNA lesions. In Arabidopsis CEN2, RAD1-RAD10 are also part of the GGR-NER repair process [25]. In order to define whether CUL4, DDB1A and DDB2 act in the same repair pathway as CEN2 and RAD1-RAD10 different double mutants were produced and characterised for their UV-C sensitivity by using the root growth assay. Double ddb1a-2 ddb2-2, cul4-1 cen2-2, ddb1a-2 cen2-2 and ddb2-2 cen2-2 mutant plants did not exhibit significant additive effects on root growth inhibition compared to the respective single mutants (Figure 3A). These results are consistent with epistatic interactions between CUL4, DDB1A, DDB2 and CEN2 indicating that they act in the same pathway.


Regulation and role of Arabidopsis CUL4-DDB1A-DDB2 in maintaining genome integrity upon UV stress.

Molinier J, Lechner E, Dumbliauskas E, Genschik P - PLoS Genet. (2008)

Genetic interactions in NER and HR.One-week-old single and double mutant plants (A) ddb1a-2-ddb2-2; cul4-1-cen2-2; ddb1a-2-cen2-2; ddb2-2-cen2-2 (B) cul4-1-rad10; ddb1a-2-rad10; ddb2-2-rad10; cen2-2-rad1 were exposed to 600 J/m2 of UV-C. Root growth was measured 24h following irradiation. Root growth was calculated relative to the corresponding untreated plants (±SEM). Eight plants per replicate were used and experiments were triplicated. For all the single and double mutants p<0.05, compared to WT plants. Because ddb2-2 is in a different Arabidopsis ecotype, the single control mutants were selected as segregants from each double mutant involving ddb2-2 and are indicated by *.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2396500&req=5

pgen-1000093-g003: Genetic interactions in NER and HR.One-week-old single and double mutant plants (A) ddb1a-2-ddb2-2; cul4-1-cen2-2; ddb1a-2-cen2-2; ddb2-2-cen2-2 (B) cul4-1-rad10; ddb1a-2-rad10; ddb2-2-rad10; cen2-2-rad1 were exposed to 600 J/m2 of UV-C. Root growth was measured 24h following irradiation. Root growth was calculated relative to the corresponding untreated plants (±SEM). Eight plants per replicate were used and experiments were triplicated. For all the single and double mutants p<0.05, compared to WT plants. Because ddb2-2 is in a different Arabidopsis ecotype, the single control mutants were selected as segregants from each double mutant involving ddb2-2 and are indicated by *.
Mentions: CENTRIN2 (CEN2) and the heterodimer RAD1-RAD10 (XPF-ERCC1) act in the GGR-NER repair pathway [7]. CEN2 is part of the XPC-RAD23 recognition complex, whereas, RAD1-RAD10 in cooperation with RAD2 are the endonucleases excising the bulky DNA lesions. In Arabidopsis CEN2, RAD1-RAD10 are also part of the GGR-NER repair process [25]. In order to define whether CUL4, DDB1A and DDB2 act in the same repair pathway as CEN2 and RAD1-RAD10 different double mutants were produced and characterised for their UV-C sensitivity by using the root growth assay. Double ddb1a-2 ddb2-2, cul4-1 cen2-2, ddb1a-2 cen2-2 and ddb2-2 cen2-2 mutant plants did not exhibit significant additive effects on root growth inhibition compared to the respective single mutants (Figure 3A). These results are consistent with epistatic interactions between CUL4, DDB1A, DDB2 and CEN2 indicating that they act in the same pathway.

Bottom Line: Moreover, we provide evidences for crosstalks between GGR, the plant-specific photo reactivation pathway and the RAD1-RAD10 endonucleases upon UV exposure.Finally, we report that DDB2 degradation upon UV stress depends not only on CUL4, but also on the checkpoint protein kinase Ataxia telangiectasia and Rad3-related (ATR).Interestingly, we found that DDB1A shuttles from the cytoplasm to the nucleus in an ATR-dependent manner, highlighting an upstream level of control and a novel mechanism of regulation of this E3 ligase.

View Article: PubMed Central - PubMed

Affiliation: Institut de Biologie Moléculaire des Plantes du CNRS (UPR2357), conventionné avec l'Université Louis Pasteur, Strasbourg, France.

ABSTRACT
Plants use the energy in sunlight for photosynthesis, but as a consequence are exposed to the toxic effect of UV radiation especially on DNA. The UV-induced lesions on DNA affect both transcription and replication and can also have mutagenic consequences. Here we investigated the regulation and the function of the recently described CUL4-DDB1-DDB2 E3 ligase in the maintenance of genome integrity upon UV-stress using the model plant Arabidopsis. Physiological, biochemical, and genetic evidences indicate that this protein complex is involved in global genome repair (GGR) of UV-induced DNA lesions. Moreover, we provide evidences for crosstalks between GGR, the plant-specific photo reactivation pathway and the RAD1-RAD10 endonucleases upon UV exposure. Finally, we report that DDB2 degradation upon UV stress depends not only on CUL4, but also on the checkpoint protein kinase Ataxia telangiectasia and Rad3-related (ATR). Interestingly, we found that DDB1A shuttles from the cytoplasm to the nucleus in an ATR-dependent manner, highlighting an upstream level of control and a novel mechanism of regulation of this E3 ligase.

Show MeSH
Related in: MedlinePlus