Chemotherapeutic approaches to anticancer treatment ideally target specific defects of transformed cells with the goal of halting their proliferation, or eliciting programmed death. Genetic approaches to the development of anticancer drugs focus on the identification secondary drug targets, or pathways, whose inhibition will result in synergistic enhancement of primary defects, and subsequent death of .cancer cells. This proposal provides evidence that a specific step in the ribosome biogenesis pathway may provide such a secondary drug target. A genome wide screen in the yeast Saccharomyces cerevisiae for mutations conferring hypersensitivity to the anticancer, antimetabolite 5-fluorouracil (SFU) identified the gene for thymidylate synthetase (CDC21), the drug's known target, as well as six genes whose products play an essential role in the production of 60S ribosomes. Four of these genes encode riboexonuclease components of the exosome, a complex of proteins required for a specific step in the processing of the 5.8S rRNA. Significantly, treatment of yeast and human cells with 5- fluorouracil results in inhibition of the same 5.8S rRNA processing step catalyzed by these enzymes, and it appears to cause cell cycle arrest in yeast. The research proposed here focuses on understanding the mechanism of 5FU inhibition of rRNA processing and the relationship of this effect to inhibition of cell cycle progression. First, the ability of 5FU to inhibit protein synthesis will be analyzed and the relationship between CDC21 and rRNA processing genes will be studied in a context that will reveal synergistic effects on cell growth and cell cycle progression. Second, the effect of 5FU on the levels of mRNAs that are regulated by the exosome will be determined to establish whether the drug affects the ability of the exosome to degrade mRNAs in the nucleus. Third, a novel gene of unknown function, that was identified as a potential 5FU target, will be characterized to determine its function in yeast and to understand what role it plays in 5FU toxicity. Finally, experiments will be carried out in human cells to determine if the antiproliferative effects of 5FU result from inhibition of 5.8S rRNA processing. These experiments should clarify the connection between 5FU and rRNA processing and therefore provide a new pathway to which anticancer drugs can be targeted.
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| Kuai, Letian; Fang, Feng; Butler, J Scott et al. (2004) Polyadenylation of rRNA in Saccharomyces cerevisiae. Proc Natl Acad Sci U S A 101:8581-6 |
