Heme deficiency sensitizes yeast cells to oxidative stress induced by hydroxyurea

Abstract

Edited by Patrick SungHydroxyurea (HU) has a long history of clinical and scientific use as an antiviral, antibacterial, and antitumor agent. It inhibits ribonucleotide reductase and reversibly arrests cells in S phase. However, high concentrations or prolonged treatment with low doses of HU can cause cell lethality. Although the cytotoxicity of HU may significantly contribute to its therapeutic effects, the underlying mechanisms remain poorly understood. We have previously shown that HU can induce cytokinesis arrest in the erg11-1 mutant of fission yeast, which has a partial defect in the biosynthesis of fungal membrane sterol ergosterol. Here, we report the identification of a new mutant in heme biosynthesis, hem13-1, that is hypersensitive to HU. We found that the HU hypersensitivity of the hem13-1 mutant is caused by oxidative stress and not by replication stress or a defect in cellular response to replication stress. The mutation is hypomorphic and causes heme deficiency, which likely sensitizes the cells to the HU-induced oxidative stress. Because the heme biosynthesis pathway is highly conserved in eukaryotes, this finding, as we show in our separate report, may help to expand the therapeutic spectrum of HU to additional pathological conditions.

The RecQ DNA helicase Rqh1 promotes Rad3ATRkinase signaling in the DNA replication checkpoint pathway of fission yeast

“…As mentioned above, HU arrests the metabolic mutants at G2/M, not S phase, as they are killed by HU via the mechanisms unrelated to replication stress (25,26). We then examined the cell cycle progression by flow cytometry (Fig.…”

“…Sensitivities to HU and various DNA damaging agents were determined by spot assay or in liquid medium as described in our previous studies (9,12,25,26). For the spot assay, 2 x 10 7 cells/ml of logarithmically growing S. pombe were diluted in five-fold steps and spotted in 3 µl onto YE6S plates or YE6S plates containing the drugs at indicated concentrations.…”

“…1 ×10 7 logarithmically growing cells were collected, fixed in ice-cold 70% ethanol, and then analysed by Accuri C6 flow cytometer as described in our previous studies (25,26).…”

Novel Cell-Killing Mechanisms of Hydroxyurea and the Implication toward Combination Therapy for the Treatment of Fungal Infections

“…SMP, terbinafine, and antifungal azoles significantly enhance the cell-killing effect of HU in S. pombe. We have recently reported two mutants, the hem13-1 and erg11-1 mutants, of the fission yeast S. pombe that are highly sensitive to HU (1,2). Since HU is a relatively safe drug and has been used for the treatment of various pathological conditions but not fungal infections, we were interested in testing the enhancement of its cytotoxicity with antifungal azoles that inhibit Erg11 and the heme synthesis inhibitor sampangine (SMP).…”

“…It is a watersoluble, well-tolerated, and well-absorbed small-molecule drug that can be used either intravenously or orally. We have previously shown in the fission yeast Schizosaccharomyces pombe that hypomorphic mutations in the heme or ergosterol biosynthesis pathway significantly sensitize the cells to HU (1,2). In this study, we further investigated the cell-killing effect of HU in various combinations with inhibitors of heme and ergosterol biosynthesis in S. pombe, the baker's yeast Saccharomyces cerevisiae, and the opportunistic pathogen Candida albicans.…”