The duration of nuclear extracellular signal-regulated kinase 1 and 2 signaling during cell cycle reentry distinguishes proliferation from apoptosis in response to asbestos

Our plain-English summary of The duration of nuclear extracellular signal-regulated kinase 1 and 2 signaling during cell cycle reentry distinguishes proliferation from apoptosis in response to asbestos, by Z. Yuan, D.J. Taatjes, B.T. Mossman, et al (Cancer Research, 2004).

The study

• Authors: Z. Yuan, D.J. Taatjes, B.T. Mossman, N.H. Heintz
• Journal: Cancer Research
• Published: 2004;64(18):6530-6
• Cited by: 29 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

Asbestos exposure causes activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) in lung epithelial cells, the targets of asbestos-associated lung carcinomas. The functional significance of ERK1/2 activation in pulmonary epithelial and mesothelial cells is unclear. Using serum-stimulated mouse alveolar type II epithelial cells as a model for cell cycle reentry, we show that the duration of phospho-ERK1/2 in the nucleus determines cell fate in response to crocidolite asbestos. In response to 10% serum, a proliferative stimulus, phosphorylated ERK1/2 initially accumulated in the nucleus, and reduction of nuclear phospho-ERK1/2 after 2 to 4 hours was followed by expression of cyclin D1 and S-phase entry. Low levels of asbestos (<0.5 microg/cm2) promoted S-phase entry in low (2%) serum through an epidermal growth factor receptor-dependent pathway but did not promote cell cycle progression or induce apoptosis in the presence of high (10%) serum-containing medium. Higher levels of asbestos (1.0 to 5.0 microg/cm2) prolonged the localization of phospho-ERK1/2 in the nucleus in the presence of high serum, impeded S-phase entry, and induced apoptosis in a dose-dependent manner. Immunofluorescence microscopy indicated that the duration of signaling by phospho-ERK1/2 in the nucleus was predictive of cell fate at any concentration of asbestos. After 8 hours of exposure, cells with nuclear phospho-ERK1/2 also were positive for nuclear localization of apoptosis-inducing factor (AIF), an early event in apoptosis. In contrast, asbestos-exposed cells that displayed cytoplasmic phospho-ERK1/2 at 8 hours expressed cyclin D1 and proceeded to S phase. Our studies show that prolonged localization of phospho-ERK1/2 in the nucleus is incompatible with expression of cyclin D1 and is predictive of asbestos-associated cell death by AIF, thereby providing an approach for determining cell fate in asbestos-induced tumorigenesis.

In plain English

This was a laboratory cell-culture study using mouse type II alveolar lung epithelial cells (C10). Cells arrested by 72 hours of serum deprivation were restimulated with 2% or 10% fetal bovine serum, with or without crocidolite asbestos at 0.2 to 5.0 micrograms per square centimetre. S-phase entry, cyclin D1, apoptosis markers and the location of phosphorylated ERK1/2 were measured. With 10% serum, 60 to 65% of cells entered S phase by 20 hours. With 2% serum, 25% did, and low asbestos doses (0.2 and 0.5 micrograms per square centimetre) increased S-phase entry (P less than 0.1 versus control). The EGFR inhibitor AG1478 markedly reduced S-phase entry with 2% serum alone or plus 0.2 micrograms per square centimetre of asbestos. Higher doses inhibited S-phase entry and increased apoptosis in a dose-dependent way (linear trend, P less than 0.001). Nearly all asbestos-exposed cells keeping phospho-ERK1/2 in the nucleus at 8 hours showed nuclear AIF, and such cells lacked cyclin D1. The authors concluded that the duration of nuclear phospho-ERK1/2 signaling predicted whether cells proliferated or died.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: In vitro cell-culture experiment using a mouse lung epithelial cell line
• Who was studied: Mouse type II alveolar lung epithelial cells (C10 line), synchronized in G0 by serum deprivation. In the cytometry assays, 8,000 to 10,000 cells were scanned per coverslip. Laboratory study at the University of Vermont.
• Exposure: Crocidolite asbestos (NIEHS reference sample) at 0.2 to 5.0 micrograms per square centimetre, applied with 0.2%, 2% or 10% fetal bovine serum
• Main results:
  • S-phase entry by 20 hours was 60 to 65% of cells with 10% FBS, about 1.0% with 0.2% FBS, and 25% with 2% FBS. Low asbestos doses (0.2 and 0.5 micrograms per square centimetre) added to 2% FBS promoted S-phase entry, while higher doses (2.5 and above) inhibited it.
  • The increase in S-phase entry at 0.2 and 0.5 micrograms per square centimetre in 2% FBS was reported at P less than 0.1 for treated versus control
  • At 5.0 micrograms per square centimetre asbestos, a dose that blocked about 90% of cells from entering S phase, about 80% of the cells expressing nuclear cyclin D1 still incorporated BrdUrd
  • The EGFR phosphorylation inhibitor AG1478 markedly reduced BrdUrd incorporation in response to 2% FBS or 2% FBS plus 0.2 micrograms per square centimetre asbestos. The authors took this to show that S-phase entry depended on EGFR signaling.
  • Catalase increased rather than decreased S-phase entry at low asbestos doses, and it did not rescue cells from arrest at higher doses. The authors took this to mean the proliferative effect did not appear to be due to fiber-generated reactive oxygen species.
  • In 10% FBS alone, about 5% of cells were positive for apoptosis markers at 20 hours. Asbestos produced a dose-dependent, linear increase in sub-G1 cells (P less than 0.001).
  • At 6 and 12 hours, asbestos raised phospho-ERK1/2 in a dose-dependent manner, and cyclin D1 was reduced. Phospho-pRB was reduced at 12 hours. Phospho-JNK appeared only at 12 hours and only at higher doses. Phospho-p38 rose about twofold at higher doses.
  • With 10% FBS alone, phospho-ERK1/2 was in the nucleus by 15 minutes and in the cytoplasm or membrane by 2 hours. With 5.0 micrograms per square centimetre asbestos, it persisted in the nuclei of a fraction of cells for at least 6 hours.
  • Nearly all cells with nuclear phospho-ERK1/2 at 8 hours after serum stimulation with asbestos also had nuclear AIF. No cell had nuclear AIF without nuclear phospho-ERK1/2. Nuclear AIF rose with asbestos dose (P less than 0.001).
  • Cells with nuclear phospho-ERK1/2 at 8 hours did not express cyclin D1, and adjacent cells often responded differently. At 5.0 micrograms per square centimetre every cell was in contact with numerous fibers, so fiber deposition alone did not explain cell fate.
• The authors' conclusion: The authors concluded that prolonged nuclear localization of phospho-ERK1/2 is incompatible with cyclin D1 expression and predicts asbestos-associated cell death via AIF, and that cyclin D1 marks cells destined to enter S phase
• Limitations the authors state:
  • The authors noted that flow cytometry and single-stranded DNA staining detect only late-stage apoptosis, so they cannot link signaling events to apoptosis in individual cells
  • The authors stated it is not known whether nuclear ERK1/2 relocates to the cell periphery or whether kinase activity in these locations is controlled independently
  • The authors said they do not yet understand why neighbouring cells, presumably at the same cell cycle stage, showed such different ERK1/2 responses

Topics

The National Library of Medicine indexes this study under subjects our site covers: Crocidolite.

Medical terms in this study, explained in our Lexicon: Apoptosis, Asbestos Exposure, Carcinoma, Mesothelial Cells, Crocidolite, Tumorigenesis.

Related studies

• Persistent localization of activated extracellular signal-regulated kinases (ERK1/2) is epithelial cell-specific in an inhalation model of asbestosis — A.B. Cummins, C. Palmer, et al. (2003)
• Different accumulation of activated extracellular signal-regulated kinases (ERK 1/2) and role in cell-cycle alterations by epidermal growth factor, hydrogen peroxide, or asbestos in pulmonary epithelial cells — S. Buder-Hoffmann, C. Palmer, et al. (2001)
• Asbestos inhalation induces tyrosine nitration associated with extracellular signal-regulated kinase 1/2 activation in the rat lung — A. Iwagaki, N. Choe, et al. (2003)
• Asbestos-mediated CREB phosphorylation is regulated by protein kinase A and extracellular signal-regulated kinases 1/2 — C.A. Barlow, T.F. Barrett, et al. (2007)
• Role of extracellular signal-regulated protein kinases in apoptosis by asbestos and H2O2 — L.A. Jiménez, C. Zanella, et al. (1997)
• Increased phosphorylated extracellular signal-regulated kinase immunoreactivity associated with proliferative and morphologic lung alterations after chrysotile asbestos inhalation in mice — R.F. Robledo, S.A. Buder-Hoffmann, et al. (2000)

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