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ATM regulates insulin-like growth factor 1-secretory clusterin (IGF-1-sCLU) expression that protects cells against senescence.

Luo X, Suzuki M, Ghandhi SA, Amundson SA, Boothman DA - PLoS ONE (2014)

Bottom Line: In contrast, administration of an IGF-1 inhibitor caused apoptosis of senescent cells.Thus, IGF-1 signaling is required for survival, whereas sCLU appears to protect cells from premature senescence, as IMR-90 cells with sCLU knockdown undergo senescence faster than control cells.Thus, the ATM-IGF-1-sCLU pathway protects cells from lethality and suspends senescence.

View Article: PubMed Central - PubMed

Affiliation: Departments of Pharmacology and Radiation Oncology, Laboratory of Molecular Cell Stress Responses, Program in Cell Stress and Cancer Nanomedicine, Simmons Cancer Center, University of Texas Southwestern Medical Center, Dallas, Texas, United States of America.

ABSTRACT
Downstream factors that regulate the decision between senescence and cell death have not been elucidated. Cells undergo senescence through three pathways, replicative senescence (RS), stress-induced premature senescence (SIPS) and oncogene-induced senescence. Recent studies suggest that the ataxia telangiectasia mutant (ATM) kinase is not only a key protein mediating cellular responses to DNA damage, but also regulates cellular senescence induced by telomere end exposure (in RS) or persistent DNA damage (in SIPS). Here, we show that expression of secretory clusterin (sCLU), a known pro-survival extracellular chaperone, is transcriptionally up-regulated during both RS and SIPS, but not in oncogene-induced senescence, consistent with a DNA damage-inducible mechanism. We demonstrate that ATM plays an important role in insulin-like growth factor 1 (IGF-1) expression, that in turn, regulates downstream sCLU induction during senescence. Loss of ATM activity, either by genomic mutation (ATM-deficient fibroblasts from an ataxia telangiectasia patient) or by administration of a chemical inhibitor (AAI, an inhibitor of ATM and ATR), blocks IGF-1-sCLU expression in senescent cells. Downstream, sCLU induction during senescence is mediated by IGF-1R/MAPK/Egr-1 signaling, identical to its induction after DNA damage. In contrast, administration of an IGF-1 inhibitor caused apoptosis of senescent cells. Thus, IGF-1 signaling is required for survival, whereas sCLU appears to protect cells from premature senescence, as IMR-90 cells with sCLU knockdown undergo senescence faster than control cells. Thus, the ATM-IGF-1-sCLU pathway protects cells from lethality and suspends senescence.

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sCLU expression is differently regulated in SIPS, RS and oncogene-induced senescence.(A) sCLU levels were increased during senescence in BJ, HE45 and HBEC cells. BJ, HE45 and HBEC cells were aged by continuous cell culture and senescence status determined by %SA-β-gal+ cells. (B) Forced expression of Ras12V represses sCLU expression. Young IMR-90 cells were transduced with lentiviral-mediated Ras12V expression. After cells exhibited typical senescence morphology, stained SA-β-gal+, and were growth arrested, they were harvested and sCLU levels monitored by Western blotting. Phospho-Erk-1/2 was probed to monitor the effects of Ras12V over-expression. α-Tubulin was used for a loading control. (C) IMR-90 cells cultured in 20% oxygen (O2) underwent senescence faster than cells cultured in 2% O2. The population doubling of IMR-90 cells cultured in 20% O2 decreased faster than cells cultured in 2% O2. (D) BJ cells were not affected by different O2 tensions. In contrast to IMR-90 cells, the population doublings of BJ cells were identical whether cultured in 20% or 2% O2. (E) sCLU levels increased in IMR-90 cells cultured in 20% O2 compared to levels found in IMR-90 cells cultured in 2% O2 in both early and late passage cells. (F) Young (Y), middle-aged (M), and senescent (S) IMR-90 cells were treated with IR (10 Gy). psCLU, sCLU, total Erk-1/2 (Erk) and phosphorylated Erk-1/2 (pErk) were detected by western blot analyses. For (C) and (D), representative results are presented from at least three repeat experiments with similar results.
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pone-0099983-g002: sCLU expression is differently regulated in SIPS, RS and oncogene-induced senescence.(A) sCLU levels were increased during senescence in BJ, HE45 and HBEC cells. BJ, HE45 and HBEC cells were aged by continuous cell culture and senescence status determined by %SA-β-gal+ cells. (B) Forced expression of Ras12V represses sCLU expression. Young IMR-90 cells were transduced with lentiviral-mediated Ras12V expression. After cells exhibited typical senescence morphology, stained SA-β-gal+, and were growth arrested, they were harvested and sCLU levels monitored by Western blotting. Phospho-Erk-1/2 was probed to monitor the effects of Ras12V over-expression. α-Tubulin was used for a loading control. (C) IMR-90 cells cultured in 20% oxygen (O2) underwent senescence faster than cells cultured in 2% O2. The population doubling of IMR-90 cells cultured in 20% O2 decreased faster than cells cultured in 2% O2. (D) BJ cells were not affected by different O2 tensions. In contrast to IMR-90 cells, the population doublings of BJ cells were identical whether cultured in 20% or 2% O2. (E) sCLU levels increased in IMR-90 cells cultured in 20% O2 compared to levels found in IMR-90 cells cultured in 2% O2 in both early and late passage cells. (F) Young (Y), middle-aged (M), and senescent (S) IMR-90 cells were treated with IR (10 Gy). psCLU, sCLU, total Erk-1/2 (Erk) and phosphorylated Erk-1/2 (pErk) were detected by western blot analyses. For (C) and (D), representative results are presented from at least three repeat experiments with similar results.

Mentions: To investigate whether sCLU expression increases were specific to cells undergoing SIPS, we examined other normal human diploid cells, BJ, HE49 and human bronchial epithelial cells (HBECs), that undergo RS, and are not subject to SIPS under the relatively elevated O2 levels found under normal tissue culture growth conditions. In all three cells, psCLU and sCLU levels were increased in cells undergoing RS (Figure 2A). Thus, sCLU levels are enhanced by both RS and SIPS.


ATM regulates insulin-like growth factor 1-secretory clusterin (IGF-1-sCLU) expression that protects cells against senescence.

Luo X, Suzuki M, Ghandhi SA, Amundson SA, Boothman DA - PLoS ONE (2014)

sCLU expression is differently regulated in SIPS, RS and oncogene-induced senescence.(A) sCLU levels were increased during senescence in BJ, HE45 and HBEC cells. BJ, HE45 and HBEC cells were aged by continuous cell culture and senescence status determined by %SA-β-gal+ cells. (B) Forced expression of Ras12V represses sCLU expression. Young IMR-90 cells were transduced with lentiviral-mediated Ras12V expression. After cells exhibited typical senescence morphology, stained SA-β-gal+, and were growth arrested, they were harvested and sCLU levels monitored by Western blotting. Phospho-Erk-1/2 was probed to monitor the effects of Ras12V over-expression. α-Tubulin was used for a loading control. (C) IMR-90 cells cultured in 20% oxygen (O2) underwent senescence faster than cells cultured in 2% O2. The population doubling of IMR-90 cells cultured in 20% O2 decreased faster than cells cultured in 2% O2. (D) BJ cells were not affected by different O2 tensions. In contrast to IMR-90 cells, the population doublings of BJ cells were identical whether cultured in 20% or 2% O2. (E) sCLU levels increased in IMR-90 cells cultured in 20% O2 compared to levels found in IMR-90 cells cultured in 2% O2 in both early and late passage cells. (F) Young (Y), middle-aged (M), and senescent (S) IMR-90 cells were treated with IR (10 Gy). psCLU, sCLU, total Erk-1/2 (Erk) and phosphorylated Erk-1/2 (pErk) were detected by western blot analyses. For (C) and (D), representative results are presented from at least three repeat experiments with similar results.
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Related In: Results  -  Collection

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pone-0099983-g002: sCLU expression is differently regulated in SIPS, RS and oncogene-induced senescence.(A) sCLU levels were increased during senescence in BJ, HE45 and HBEC cells. BJ, HE45 and HBEC cells were aged by continuous cell culture and senescence status determined by %SA-β-gal+ cells. (B) Forced expression of Ras12V represses sCLU expression. Young IMR-90 cells were transduced with lentiviral-mediated Ras12V expression. After cells exhibited typical senescence morphology, stained SA-β-gal+, and were growth arrested, they were harvested and sCLU levels monitored by Western blotting. Phospho-Erk-1/2 was probed to monitor the effects of Ras12V over-expression. α-Tubulin was used for a loading control. (C) IMR-90 cells cultured in 20% oxygen (O2) underwent senescence faster than cells cultured in 2% O2. The population doubling of IMR-90 cells cultured in 20% O2 decreased faster than cells cultured in 2% O2. (D) BJ cells were not affected by different O2 tensions. In contrast to IMR-90 cells, the population doublings of BJ cells were identical whether cultured in 20% or 2% O2. (E) sCLU levels increased in IMR-90 cells cultured in 20% O2 compared to levels found in IMR-90 cells cultured in 2% O2 in both early and late passage cells. (F) Young (Y), middle-aged (M), and senescent (S) IMR-90 cells were treated with IR (10 Gy). psCLU, sCLU, total Erk-1/2 (Erk) and phosphorylated Erk-1/2 (pErk) were detected by western blot analyses. For (C) and (D), representative results are presented from at least three repeat experiments with similar results.
Mentions: To investigate whether sCLU expression increases were specific to cells undergoing SIPS, we examined other normal human diploid cells, BJ, HE49 and human bronchial epithelial cells (HBECs), that undergo RS, and are not subject to SIPS under the relatively elevated O2 levels found under normal tissue culture growth conditions. In all three cells, psCLU and sCLU levels were increased in cells undergoing RS (Figure 2A). Thus, sCLU levels are enhanced by both RS and SIPS.

Bottom Line: In contrast, administration of an IGF-1 inhibitor caused apoptosis of senescent cells.Thus, IGF-1 signaling is required for survival, whereas sCLU appears to protect cells from premature senescence, as IMR-90 cells with sCLU knockdown undergo senescence faster than control cells.Thus, the ATM-IGF-1-sCLU pathway protects cells from lethality and suspends senescence.

View Article: PubMed Central - PubMed

Affiliation: Departments of Pharmacology and Radiation Oncology, Laboratory of Molecular Cell Stress Responses, Program in Cell Stress and Cancer Nanomedicine, Simmons Cancer Center, University of Texas Southwestern Medical Center, Dallas, Texas, United States of America.

ABSTRACT
Downstream factors that regulate the decision between senescence and cell death have not been elucidated. Cells undergo senescence through three pathways, replicative senescence (RS), stress-induced premature senescence (SIPS) and oncogene-induced senescence. Recent studies suggest that the ataxia telangiectasia mutant (ATM) kinase is not only a key protein mediating cellular responses to DNA damage, but also regulates cellular senescence induced by telomere end exposure (in RS) or persistent DNA damage (in SIPS). Here, we show that expression of secretory clusterin (sCLU), a known pro-survival extracellular chaperone, is transcriptionally up-regulated during both RS and SIPS, but not in oncogene-induced senescence, consistent with a DNA damage-inducible mechanism. We demonstrate that ATM plays an important role in insulin-like growth factor 1 (IGF-1) expression, that in turn, regulates downstream sCLU induction during senescence. Loss of ATM activity, either by genomic mutation (ATM-deficient fibroblasts from an ataxia telangiectasia patient) or by administration of a chemical inhibitor (AAI, an inhibitor of ATM and ATR), blocks IGF-1-sCLU expression in senescent cells. Downstream, sCLU induction during senescence is mediated by IGF-1R/MAPK/Egr-1 signaling, identical to its induction after DNA damage. In contrast, administration of an IGF-1 inhibitor caused apoptosis of senescent cells. Thus, IGF-1 signaling is required for survival, whereas sCLU appears to protect cells from premature senescence, as IMR-90 cells with sCLU knockdown undergo senescence faster than control cells. Thus, the ATM-IGF-1-sCLU pathway protects cells from lethality and suspends senescence.

Show MeSH
Related in: MedlinePlus