5-AZACYTIDINE TREATMENT OF THE FISSION YEAST LEADS TO CYTOTOXICITY AND CELL-CYCLE ARREST

A fission yeast gene which shares considerable sequence homology with cytosine-specific DNA methyltransferases has recently been identified. This discovery has led us to investigate the effects of the treatment of fission yeast with the nucleoside analogue 5-azacytidine (5-azaC). 5-AzaC is known to inhibit cytosine methylation as a result of the fo rmation of stable covalent complexes between DNA (cytosine-5) methyltr ansferases (C5 Mtases) and 5-azaC containing DNA. Here we demonstrate that 5-azaC treatment of Schizosaccharomyces pombe leads to reversible cell cycle arrest at the G2/M transition. This reversible arrest is d ependent on the cell cycle checkpoint mechanisms which act to prevent the onset of mitosis in the presence of either damaged or unreplicated DNA. Treatment of S. pombe cell division cycle and checkpoint mutants indicates that 5-azaC causes DNA damage and is likely to inhibit a la te stage in DNA replication. The data show that viability in the prese nce of the drug requires both the DNA damage and the replication check point pathways to be functional. 5-AzaC also elicits a transcriptional response which is associated with DNA damage and the inhibition of DN A replication in fission yeast, and this response is absent in cells c arrying G2 checkpoint mutations. The implications of these observation s for both the use of 5-azaC in cancer chemotherapy and the existence of cytosine methylation in fission yeast are discussed.