9514196 Langmore Telomeres are the terminal nucleoproteins of all eukaryotic chromosomes. The definitive function of telomeres is stabilization of the ends of chromosomes, apparently by protection against non-homologous recombination and exonucleolytic degradation--well-documented instabilities of non-telomere ends in cells and cell extracts. Telomeres must maintain this protective function to prevent degradation, rearrangement, or loss of chromosomes. Deterioration of telomere function might be involved in some of the chromosomal instabilities associated with developmental defects and aging. Genetic and biochemical studies of the highly diverse, short, telomeres in protozoa and fungi have probed telomere function in unicellular, immortal organisms. Vertebrates telomeres have a highly conserved repeating sequence (TTAGGG)n of length 2-100 kb, unusual nucleosome structure, and probably specific non-histone proteins. The role of these components in the protective function of vertebrate telomeres is not clear due to the difficulties in doing genetic studies, and the lack of an vitro model for telomere assembly and behavior. The ideal model system will permit the protective function of telomeres to be studied independent of the possible roles of telomeres in the cell cycle, chromosome segregation, and gene regulation. The aim of this project is to understand how telomeres are protected, using a reductionist approach of recapitulating and manipulating telomere structure and function in vitro. Xenopus egg extracts are used for this task, because they have both the strong non-homologous recombination activity and specific telomere-protection activity characteristic of human and other vertebrate somatic cells. The specific goals are to: 1) Describe model telomere assembly and function in vitro and in vivo; and 2) Determine the DNA structure requirements for model telomere stability. These studies will help to understand telomere roles in chromosome and cell behavior. The in vitro studi es will lead to new hypotheses about the mechanism of chromosome stability and instability that will be useful in explaining chromosome behavior, and for suggesting new experiments. The identification and characterization of the factors that are necessary and sufficient for protection of telomere ends from recombination will lead to even more detailed mechanistic and structural studies. ***
