Redox regulation of the G1 to S phase transition in the mouse embryo fibroblast cell cycle

Sarita G Menon; Ehab H Sarsour; Douglas R Spitz; Ryuji Higashikubo; Mary Sturm; Hannah Zhang; Prabhat C Goswami

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Redox regulation of the G1 to S phase transition in the mouse embryo fibroblast cell cycle

Journal article

Peer reviewed

Redox regulation of the G1 to S phase transition in the mouse embryo fibroblast cell cycle

Sarita G Menon, Ehab H Sarsour, Douglas R Spitz, Ryuji Higashikubo, Mary Sturm, Hannah Zhang and Prabhat C Goswami

Cancer research (Chicago, Ill.), Vol.63(9), pp.2109-2117

05/01/2003

PMID: 12727827

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Abstract

The hypothesis that intracellular oxidation/reduction (redox) reactions regulate the G(0)-G(1) to S-phase transition in the mouse embryonic fibroblast cell cycle was investigated. Intracellular redox state was modulated with a thiol-antioxidant, N-acetyl-L-cysteine (NAC), and cell cycle progression was measured using BrdUrd pulse-chase and flow cytometric analysis. Treatment with NAC for 12 h resulted in an approximately 6-fold increase in intracellular low-molecular-weight thiols and a decrease in the MFI of an oxidation-sensitive probe, dihydrofluorescein diacetate, indicating a shift in the intracellular redox state toward a more reducing environment. NAC-induced alterations in redox state caused selective delays in progression from G(0)-G(1) to S phase in serum-starved cells that were serum stimulated to reenter the cell cycle as well as to inhibit progression from G(1) to S phase in asynchronous cultures with no significant alterations in S phase, and G(2)+M transits. NAC treatment also showed a 70% decrease in cyclin D1 protein levels and a 3-4-fold increase in p27 protein levels, which correlated with decreased retinoblastoma protein phosphorylation. Cells released from the NAC treatment showed a transient increase in dihydrofluorescein fluorescence and oxidized glutathione content between 0 and 8 h after release, indicating a shift in intracellular redox state to a more oxidizing environment. These changes in redox state were followed by an increase in cyclin D1, a decrease in p27, retinoblastoma protein hyperphosphorylation and subsequent entry into S phase by 8-12 h after the removal of NAC. These results support the hypothesis that a redox cycle within the mammalian cell cycle might provide a mechanistic link between the metabolic processes early in G(1) and the activation of G(1)-regulatory proteins in preparation for the entry of cells into S phase.

Oxidation-Reduction

Embryo, Mammalian

Cell Cycle Proteins - metabolism

Antioxidants - pharmacology

G1 Phase - drug effects

G1 Phase - physiology

Cell Cycle Proteins - antagonists & inhibitors

Tumor Suppressor Protein p53 - physiology

Animals

Flow Cytometry

Signal Transduction - drug effects

Acetylcysteine - pharmacology

S Phase - physiology

Signal Transduction - physiology

Fibroblasts - cytology

Mice

S Phase - drug effects

Fibroblasts - metabolism

Details

Title: Subtitle: Redox regulation of the G1 to S phase transition in the mouse embryo fibroblast cell cycle
Creators: Sarita G Menon - Free Radical and Radiation Biology Program, Department of Radiation Oncology, University of Iowa, Iowa City 52242, USA
Ehab H Sarsour
Douglas R Spitz
Ryuji Higashikubo
Mary Sturm
Hannah Zhang
Prabhat C Goswami
Resource Type: Journal article
Publication Details: Cancer research (Chicago, Ill.), Vol.63(9), pp.2109-2117
Publisher: United States
PMID: 12727827
ISSN: 0008-5472
eISSN: 1538-7445
Grant note: CA66081 / NCI NIH HHS HL51469 / NHLBI NIH HHS CA69593 / NCI NIH HHS
Language: English
Date published: 05/01/2003
Academic Unit: Pathology; Radiation Oncology
Record Identifier: 9984046806102771

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