Telomeres are the nucleoprotein complexes that allow the complete replication of chromosome ends and distinguish these ends from double-strand breaks. Telomere DNA consists of tandem arrays of TG-rich repeats. The length of these repeat tracts is kept nearly constant in human germ cells and yeast by regulating the processes of lengthening, via telomerase, and shortening, due to incomplete replication or nucleolytic degradation. How these two processes are regulated to give a constant telomere length is unknown. In human somatic cells, the length of the repeats decreases as cells divide, leading to cell senescence when telomeres become too short. How telomere length information is transmitted to the cell cycle machinery is unknown. Our long-term goal is to use yeast as a model system to understand these processes. Yeast measure telomere length by counting the number of molecules of the major telomere binding protein Raplp; however, how Raplp molecules are counted is under debate. We developed a model for telomere length regulation based on our construction of yeast synthetic telomeres. We propose that yeast telomeres form a folded structure with Raplp and 2 regulatory proteins, Riflp and Rif2p, to count Raplp molecules and block elongation. Short telomeres have too few Raplp molecules to form this structure, so they are elongated. Other models suggest that Sir proteins also regulate length, and we have devised assays to directly test this possibility. Tel2p binds to telomeres in vivo and regulates their length. Tel2p mutants alter telomere length and the cellular response to a single double-strand break and other forms of DNA damage. We hypothesize that Tel2p binds to telomeres and double-strand breaks to regulate activation of DNA damage checkpoints. The activities of the checkpoint proteins Tellp and Meclp suggest that they are involved in these processes.
Our specific aims are to determine 1) the protein composition of the structure that regulates telomere length; 2) if Tellp and Meclp are recruited to telomeres of different lengths; and 3) how Tel2p alters telomere length and the cellular response to DNA damage and if Tel2p acts at double-strand breaks in vivo.
Showing the most recent 10 out of 20 publications
