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Phosphorylation of SMC1 by ATR is required for desferrioxamine (DFO)-induced apoptosis.

So EY, Ausman M, Saeki T, Ouchi T - Cell Death Dis (2011)

Bottom Line: Among these sites, phosphorylation of SMC1, NBS1, and Chk1 by DFO are mediated by ATR as it is greatly reduced in both ATR-deficient human fibroblasts and HCT116 human colon cancer cells in which ATR is heterozygously mutated, whereas these proteins are phosphorylated in cells deficient for ATM and DNA-PKcs.DFO-induced apoptosis is decreased in ATR-mutant HCT116 cells, although p53 is normally activated in those cells.These results suggest that DFO induces apoptosis through the ATR-SMC1 arm of the pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, NUHS, Systems Biology Program, Pritzker School of Medicine, The University of Chicago, Evanston, IL 60201, USA.

ABSTRACT
DNA damage signaling pathways are initiated in response to chemical reagents and radiation damage, as well as in response to hypoxia. It is implicated that structural maintenance of chromosomes 1 (SMC1) is not only a component of the cohesion complex but also facilitates the activation of DNA damage checkpoint proteins. Here, we studied the mechanism of DNA damage checkpoint activated by ATR-SMC1 pathway when cells are treated with desferrioxamine (DFO), a hypoxia-mimetic reagent. We show that DFO treatment induces phosphorylation of SMC1 at Ser966, NBS1 at Ser343, Chk1 at Ser317, Chk2 at Thr68, and p53 at Ser15. Among these sites, phosphorylation of SMC1, NBS1, and Chk1 by DFO are mediated by ATR as it is greatly reduced in both ATR-deficient human fibroblasts and HCT116 human colon cancer cells in which ATR is heterozygously mutated, whereas these proteins are phosphorylated in cells deficient for ATM and DNA-PKcs. DFO-induced apoptosis is decreased in ATR-mutant HCT116 cells, although p53 is normally activated in those cells. Expression of SMC1 S966A in which Ser966 is substituted to Ala attenuates apoptosis and phosphorylation of Chk1 at Ser317 after DFO treatment, although levels of HIF1α are not significantly changed. These results suggest that DFO induces apoptosis through the ATR-SMC1 arm of the pathway.

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Schematic presentation of pathways indicating SMC1 is involved in DFO-induced apoptosis
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fig7: Schematic presentation of pathways indicating SMC1 is involved in DFO-induced apoptosis

Mentions: It is shown recently that p53, SMC1, p53-binding protein (53BP1), and Chk1/2 are phosphorylated by ATR in response to IR in ATM-, Mre11-, and cell cycle-independent manners.15 As demonstrated in Figure 2, we found that DFO-induced SMC1 phosphorylation at Ser966 was significantly lowered in ATR-deficient fibroblasts, but not in ATM-deficient cells. Phosphorylation of SMC1 at Ser966 by NCS was detected in HCT116-ATR(−/flox) cells, although it is slightly weaker than the other cell lines examined. Given that this phosphorylation of SMC1 by DFO is greatly reduced in HCT116-ATR(−/flox) cells, these results implicate that ATR is critical for SMC1's phosphorylation in DFO pathway. Recent studies have illustrated that phosphorylation of SMC1 is required for the activation of S-phase checkpoint after IR, and phosphorylation of NBS1 is essential for this phosphorylation of SMC1.27 Our results showed that DFO induces strong phosphorylation of SMC1 at Ser966, although phosphorylation of NBS1 at Ser343 is quite low in wild-type ATR cells (Figure 1). Taken together, we consider that SMC1 can be activated by at least two distinct pathways, one is ATM/NBS1/SMC1 in response to IR and the other is ATR/SMC1 in response to DFO (Figure 7).


Phosphorylation of SMC1 by ATR is required for desferrioxamine (DFO)-induced apoptosis.

So EY, Ausman M, Saeki T, Ouchi T - Cell Death Dis (2011)

Schematic presentation of pathways indicating SMC1 is involved in DFO-induced apoptosis
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3101822&req=5

fig7: Schematic presentation of pathways indicating SMC1 is involved in DFO-induced apoptosis
Mentions: It is shown recently that p53, SMC1, p53-binding protein (53BP1), and Chk1/2 are phosphorylated by ATR in response to IR in ATM-, Mre11-, and cell cycle-independent manners.15 As demonstrated in Figure 2, we found that DFO-induced SMC1 phosphorylation at Ser966 was significantly lowered in ATR-deficient fibroblasts, but not in ATM-deficient cells. Phosphorylation of SMC1 at Ser966 by NCS was detected in HCT116-ATR(−/flox) cells, although it is slightly weaker than the other cell lines examined. Given that this phosphorylation of SMC1 by DFO is greatly reduced in HCT116-ATR(−/flox) cells, these results implicate that ATR is critical for SMC1's phosphorylation in DFO pathway. Recent studies have illustrated that phosphorylation of SMC1 is required for the activation of S-phase checkpoint after IR, and phosphorylation of NBS1 is essential for this phosphorylation of SMC1.27 Our results showed that DFO induces strong phosphorylation of SMC1 at Ser966, although phosphorylation of NBS1 at Ser343 is quite low in wild-type ATR cells (Figure 1). Taken together, we consider that SMC1 can be activated by at least two distinct pathways, one is ATM/NBS1/SMC1 in response to IR and the other is ATR/SMC1 in response to DFO (Figure 7).

Bottom Line: Among these sites, phosphorylation of SMC1, NBS1, and Chk1 by DFO are mediated by ATR as it is greatly reduced in both ATR-deficient human fibroblasts and HCT116 human colon cancer cells in which ATR is heterozygously mutated, whereas these proteins are phosphorylated in cells deficient for ATM and DNA-PKcs.DFO-induced apoptosis is decreased in ATR-mutant HCT116 cells, although p53 is normally activated in those cells.These results suggest that DFO induces apoptosis through the ATR-SMC1 arm of the pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, NUHS, Systems Biology Program, Pritzker School of Medicine, The University of Chicago, Evanston, IL 60201, USA.

ABSTRACT
DNA damage signaling pathways are initiated in response to chemical reagents and radiation damage, as well as in response to hypoxia. It is implicated that structural maintenance of chromosomes 1 (SMC1) is not only a component of the cohesion complex but also facilitates the activation of DNA damage checkpoint proteins. Here, we studied the mechanism of DNA damage checkpoint activated by ATR-SMC1 pathway when cells are treated with desferrioxamine (DFO), a hypoxia-mimetic reagent. We show that DFO treatment induces phosphorylation of SMC1 at Ser966, NBS1 at Ser343, Chk1 at Ser317, Chk2 at Thr68, and p53 at Ser15. Among these sites, phosphorylation of SMC1, NBS1, and Chk1 by DFO are mediated by ATR as it is greatly reduced in both ATR-deficient human fibroblasts and HCT116 human colon cancer cells in which ATR is heterozygously mutated, whereas these proteins are phosphorylated in cells deficient for ATM and DNA-PKcs. DFO-induced apoptosis is decreased in ATR-mutant HCT116 cells, although p53 is normally activated in those cells. Expression of SMC1 S966A in which Ser966 is substituted to Ala attenuates apoptosis and phosphorylation of Chk1 at Ser317 after DFO treatment, although levels of HIF1α are not significantly changed. These results suggest that DFO induces apoptosis through the ATR-SMC1 arm of the pathway.

Show MeSH
Related in: MedlinePlus