Telomeres protect chromosome ends in eukaryotes. In the absence of telomerase, telomeres shorten, which eventually leads to senescence. Alternative lengthening of telomeres (ALT) is a recombination pathway that maintains eukaryotic telomeres in cells lacking telomerase. Notably, 15% of cancers take advantage of ALT to maintain their telomeres. Also, the anti-tumor effect that telomerase-inhibiting drugs have on some cancers is compromised due to activation of ALT. However, despite the relevance of ALT to cancer development and pro- gression, its mechanism remains elusive. In large part, this is because quantitative assessment of ALT is hin- dered by its highly stochastic nature. The focus of this research is to overcome this problem by combining ex- perimental molecular genetics with a modeling approach we call Population Genetic based Modeling (PGM). This approach not only accommodates, but actually exploits the stochasticity of ALT to enable quantitative as- sessments and modeling of the ALT mechanism. Due to the lack of convenient experimental system in mam- mals, studies will be carried out in yeast Saccharomyces cerevisiae, which previously proved to be a dependable and productive model system to characterize the underlying mechanism of ALT (break-induced replication (BIR)), to define genetic requirements, and to characterize outcomes of ALT. The first goal of the project is to model the process of telomere erosion in large yeast populations, which will be accomplished by a combination of molecular genetics and PGM. The resulting models will characterize the entire population?s telomere dynamics and will allow detection of even small cell subpopulations, including putative ALT precursors (cells that eventually give rise to ALT survivors). Secondly, a similar approach will provide the first quantitative assessment of ALT frequency. Finally, the connection between the pattern of telomere erosion and ALT survivor formation will be elucidated by assessing the effects of various genetic factors and environmental stressors on both telomere erosion and ALT survivor formation. In particular, the effect of mutations affecting the DNA damage-induced checkpoints, telomere capping, and DNA repair (with an emphasis on BIR genes) on the dynamics of telomere erosion and ALT survivor formation will be evaluated. Also, the effect of various environmental stressors, includ- ing ethanol, caffeine, as well as cadmium and arsenic, known to affect telomere length, will be assessed. To- gether, the proposed research will link the formation of ALT survivors with the mode of telomere erosion and will create a comprehensive and quantitative model of ALT, which will generate new hypotheses that will guide future research aimed at unraveling the entire mechanism of ALT progression. In addition, the results of this research will provide an opportunity to use telomere erosion and ALT formation as a new type of biosensor that can be used to assess the effects of various environmental assaults on genetic stability. Finally, it is expected that the approaches and results produced through this research in yeast will be applied in the future towards elucidating ALT in humans, and particularly its roles in cancer.
Alternative lengthening of telomeres (ALT) is responsible for telomere maintenance in some cancers and can be activated in other cancers by anti-telomerase treatments. The proposed research will unravel the mechanism of ALT by linking the emergence of ALT survivors to the early precursor cells that arise during the process of telomere erosion. This work will also characterize the frequency of ALT in cell populations and the effects of various mutant backgrounds as well as of environmental stressors on ALT.
