Disruption of Yeast Forkhead-associated Cell Cycle Transcription by Oxidative Stress

Abstract

The effects of oxidative stress on yeast cell cycle depend on the stress-exerting agent. We studied the effects of two oxidative stress agents, hydrogen peroxide (HP) and the superoxide-generating agent menadione (MD). We found that two small coexpressed groups of genes regulated by the Mcm1-Fkh2-Ndd1 transcription regulatory complex are sufficient to account for the difference in the effects of HP and MD on the progress of the cell cycle, namely, G1 arrest with MD and an S phase delay followed by a G2/M arrest with HP. Support for this hypothesis is provided by fkh1fkh2 double mutants, which are affected by MD as we find HP affects wild-type cells. The apparent involvement of a forkhead protein in HP-induced cell cycle arrest, similar to that reported for Caenorhabditis elegans and human, describes a potentially novel stress response pathway in yeast. INTRODUCTIONReactive oxygen species (ROS) are by-products of aerobic metabolism, but are also attributes of the extracellular environment. They pose a threat to organisms by damaging a variety of cellular macromolecules, including DNA, membrane lipids, and proteins and are implicated with carcinogenesis, aging, and numerous degenerative diseases (Finkel and Holbrook, 2000;Neumann et al., 2003). The major ROS derived from oxygen are superoxide ions, hydrogen peroxide (HP), and hydroxy radicals, ordered here by increasing reactivity (reviewed in Shackelford et al., 2000).Oxidative stress in yeast has been studied by exposing cells to agents that are already reactive, typically HP, or drugs that cause the intracellular accumulation of reactive oxygen species (Godon et al., 1998;Dumond et al., 2000;Gasch et al., 2000). Menadione (MD) is such a drug, generating reactive superoxide ions, which can be further oxidized to give HP (Monks et al., 1992;Shackelford et al., 2000). It has been previously reported that exposure to HP or MD results in a cell cycle arrest (Flattery-O'Brien and Dawes, 1998;Leroy et al., 2001). However, the arrest points are not similar for the two agents. MD was reported to arrest cells at the G1 phase of the cell cycle, whereas HP was suggested to cause a G2 arrest by an alternate mechanism from that affected by MD (Flattery-O'Brien and Dawes, 1998); others reported that HP exposure causes a delay in the S phase (Leroy et al., 2001).Hundreds of genes are regulated so that they are expressed at specific times in the cell cycle and not at others (Cho et al., 1998;Spellman et al., 1998). The major part of this coordination is achieved by the sequential activation of a small number of transcription regulators (reviewed in Mendenhall and Hodge, 1998): MBF (a complex of Mbp1p and Swi6p) and SBF (a complex of Swi4p and Swi6p) are responsible for activating G1 transcription (Koch et al., 1993); Fkh1p and a complex formed by the cooperative promoter binding of Mcm1p and Fkh2p and a later recruitment of Ndd1p are responsible for activating G2/M transcription (Koranda et al., 2000;Kumar et al., 2000;Pic et al., 2000;Hollenhorst et al., 2001); Swi5p and Ace2p...

Reverse engineering dynamic temporal models of biological processes and their relationships

“…5). Segments I, II, and IV are the time frames that cell cycle analyses indicated the G1, S, G2/M phases predominated (27). Also note that GOALIE accurately determined the length of peroxide treatment.…”

“…Cell cycle analysis of this segment indicated that the percent contribution of each phase of the cell cycle were approximately the same. MD treatment results in G1 arrest (27), and the segmentation obtained by GOALIE corresponded to the G1, S, G2/M, and G1 phases of the cell cycle (four segments), with accurate identification of entry into G1 arrest (Fig. 5).…”

“…The GOALIE framework is validated through analysis of five yeast gene expression datasets, including two YMC time courses involving two different strains grown under two different conditions (YMC1: CEN.PK122 diploid strain, glucose-limited cultures (5) and YMC2: IFO 0233 diploid strain, not glucose limited (7)), a YCC dataset after release from α-factor synchronization (YCC: DBY8724 strain (6)), and observations of the cell cycle under treatment of (HP (27) and (MD (27).…”

“…The effects of HP and MD on yeast strain DBY8724 were evaluated recently through temporal transcriptional profiling (27). In the case of the peroxide treatment, cells were synchronized with α-factor, exposed to HP for a set period of time, and subsequently released from the oxidative stress.…”

Quantifying fitness distributions and phenotypic relationships in recombinant yeast populations

“…Briefly, H 2 O 2 (15) and menadione (16) are both inducers of oxidative stress, although their specific mechanisms of action vary. SP600125 (17) is a putative Jun N-terminal kinase (JNK) inhibitor, and diphenyleneiodonium (18) is a flavoprotein inhibitor.…”