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DNA damage shifts circadian clock time via Hausp-dependent Cry1 stabilization.

Papp SJ, Huber AL, Jordan SD, Kriebs A, Nguyen M, Moresco JJ, Yates JR, Lamia KA - Elife (2015)

Bottom Line: We demonstrate that genotoxic stress stimulates Cry1 phosphorylation and its deubiquitination by Herpes virus associated ubiquitin-specific protease (Hausp, a.k.a Usp7), stabilizing Cry1 and shifting circadian clock time.Indeed, the transcriptional response to genotoxic stress is enhanced in Cry1-/- and blunted in Cry2-/- cells.Furthermore, Cry2-/- cells accumulate damaged DNA.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemical Physiology, Scripps Research Institute, La Jolla, United States.

ABSTRACT
The circadian transcriptional repressors cryptochrome 1 (Cry1) and 2 (Cry2) evolved from photolyases, bacterial light-activated DNA repair enzymes. In this study, we report that while they have lost DNA repair activity, Cry1/2 adapted to protect genomic integrity by responding to DNA damage through posttranslational modification and coordinating the downstream transcriptional response. We demonstrate that genotoxic stress stimulates Cry1 phosphorylation and its deubiquitination by Herpes virus associated ubiquitin-specific protease (Hausp, a.k.a Usp7), stabilizing Cry1 and shifting circadian clock time. DNA damage also increases Cry2 interaction with Fbxl3, destabilizing Cry2. Thus, genotoxic stress increases the Cry1/Cry2 ratio, suggesting distinct functions for Cry1 and Cry2 following DNA damage. Indeed, the transcriptional response to genotoxic stress is enhanced in Cry1-/- and blunted in Cry2-/- cells. Furthermore, Cry2-/- cells accumulate damaged DNA. These results suggest that Cry1 and Cry2, which evolved from DNA repair enzymes, protect genomic integrity via coordinated transcriptional regulation.

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Related in: MedlinePlus

Circadian time of exposure determines phase shift in response to DNA damage.Typical results of continuous monitoring of luciferase activity from mouse embryonic fibroblasts expressing Per2::Luciferase treated with 0 (black curves) or 10 Gy (red curves) ionizing radiation 4 (CT4) or 18 (CT18) hours after circadian synchronization with 1 μM dexamethasone. Data represent the mean luciferase counts of six samples per condition from one of two independent experiments. Right: quantitation of the differences in initial circadian phase of luciferase activity caused by irradiation. Data represent the mean ± propagated s.d. difference between initial phase in Mock vs irradiated samples for six samples per condition. ***p < 0.001 for a significant interaction between CT and irradiation by 2-way ANOVA.DOI:http://dx.doi.org/10.7554/eLife.04883.013
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fig3s3: Circadian time of exposure determines phase shift in response to DNA damage.Typical results of continuous monitoring of luciferase activity from mouse embryonic fibroblasts expressing Per2::Luciferase treated with 0 (black curves) or 10 Gy (red curves) ionizing radiation 4 (CT4) or 18 (CT18) hours after circadian synchronization with 1 μM dexamethasone. Data represent the mean luciferase counts of six samples per condition from one of two independent experiments. Right: quantitation of the differences in initial circadian phase of luciferase activity caused by irradiation. Data represent the mean ± propagated s.d. difference between initial phase in Mock vs irradiated samples for six samples per condition. ***p < 0.001 for a significant interaction between CT and irradiation by 2-way ANOVA.DOI:http://dx.doi.org/10.7554/eLife.04883.013

Mentions: It has been reported that DNA damage causes phase shifts of circadian rhythms (Oklejewicz et al., 2008; Engelen et al., 2013). Consistently, we observed phase shifts in primary MEFs with a peak shift following irradiation at CT2-4, (Figure 3—figure supplement 3). The requirement for Hausp in stabilization of nuclear Cry1 after DNA damage suggested Hausp could contribute to phase shifts in response to DNA damage. By examining the circadian phase of control and Hausp-depleted fibroblasts after exposure to irradiation at CT3, we found that although the circadian phase of the non-irradiated cells is similar (Figure 2D), DNA damage-induced phase shifts were greatly diminished in Hausp-deficient fibroblasts (Figure 3D–F).


DNA damage shifts circadian clock time via Hausp-dependent Cry1 stabilization.

Papp SJ, Huber AL, Jordan SD, Kriebs A, Nguyen M, Moresco JJ, Yates JR, Lamia KA - Elife (2015)

Circadian time of exposure determines phase shift in response to DNA damage.Typical results of continuous monitoring of luciferase activity from mouse embryonic fibroblasts expressing Per2::Luciferase treated with 0 (black curves) or 10 Gy (red curves) ionizing radiation 4 (CT4) or 18 (CT18) hours after circadian synchronization with 1 μM dexamethasone. Data represent the mean luciferase counts of six samples per condition from one of two independent experiments. Right: quantitation of the differences in initial circadian phase of luciferase activity caused by irradiation. Data represent the mean ± propagated s.d. difference between initial phase in Mock vs irradiated samples for six samples per condition. ***p < 0.001 for a significant interaction between CT and irradiation by 2-way ANOVA.DOI:http://dx.doi.org/10.7554/eLife.04883.013
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4352707&req=5

fig3s3: Circadian time of exposure determines phase shift in response to DNA damage.Typical results of continuous monitoring of luciferase activity from mouse embryonic fibroblasts expressing Per2::Luciferase treated with 0 (black curves) or 10 Gy (red curves) ionizing radiation 4 (CT4) or 18 (CT18) hours after circadian synchronization with 1 μM dexamethasone. Data represent the mean luciferase counts of six samples per condition from one of two independent experiments. Right: quantitation of the differences in initial circadian phase of luciferase activity caused by irradiation. Data represent the mean ± propagated s.d. difference between initial phase in Mock vs irradiated samples for six samples per condition. ***p < 0.001 for a significant interaction between CT and irradiation by 2-way ANOVA.DOI:http://dx.doi.org/10.7554/eLife.04883.013
Mentions: It has been reported that DNA damage causes phase shifts of circadian rhythms (Oklejewicz et al., 2008; Engelen et al., 2013). Consistently, we observed phase shifts in primary MEFs with a peak shift following irradiation at CT2-4, (Figure 3—figure supplement 3). The requirement for Hausp in stabilization of nuclear Cry1 after DNA damage suggested Hausp could contribute to phase shifts in response to DNA damage. By examining the circadian phase of control and Hausp-depleted fibroblasts after exposure to irradiation at CT3, we found that although the circadian phase of the non-irradiated cells is similar (Figure 2D), DNA damage-induced phase shifts were greatly diminished in Hausp-deficient fibroblasts (Figure 3D–F).

Bottom Line: We demonstrate that genotoxic stress stimulates Cry1 phosphorylation and its deubiquitination by Herpes virus associated ubiquitin-specific protease (Hausp, a.k.a Usp7), stabilizing Cry1 and shifting circadian clock time.Indeed, the transcriptional response to genotoxic stress is enhanced in Cry1-/- and blunted in Cry2-/- cells.Furthermore, Cry2-/- cells accumulate damaged DNA.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemical Physiology, Scripps Research Institute, La Jolla, United States.

ABSTRACT
The circadian transcriptional repressors cryptochrome 1 (Cry1) and 2 (Cry2) evolved from photolyases, bacterial light-activated DNA repair enzymes. In this study, we report that while they have lost DNA repair activity, Cry1/2 adapted to protect genomic integrity by responding to DNA damage through posttranslational modification and coordinating the downstream transcriptional response. We demonstrate that genotoxic stress stimulates Cry1 phosphorylation and its deubiquitination by Herpes virus associated ubiquitin-specific protease (Hausp, a.k.a Usp7), stabilizing Cry1 and shifting circadian clock time. DNA damage also increases Cry2 interaction with Fbxl3, destabilizing Cry2. Thus, genotoxic stress increases the Cry1/Cry2 ratio, suggesting distinct functions for Cry1 and Cry2 following DNA damage. Indeed, the transcriptional response to genotoxic stress is enhanced in Cry1-/- and blunted in Cry2-/- cells. Furthermore, Cry2-/- cells accumulate damaged DNA. These results suggest that Cry1 and Cry2, which evolved from DNA repair enzymes, protect genomic integrity via coordinated transcriptional regulation.

Show MeSH
Related in: MedlinePlus