Telomere Structure In Drosophila

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We are interested in the genetic control of telomere structure and identity. Mutations are known that drastically increase or decrease the frequency of retrotransposon addition to a chromosome end, suggesting that this process is under genetic control. We have identified one gene, Telomere elongation, which has mutations that increase the frequency of terminal gene conversion, and are using positional information to clone the gene. Sequencing of two mutant alleles suggests the mutant produces a truncated form of a DNA repair protein. Chromatin structure also regulates telomere length by controlling transcription of the telomeric retrotransposons and by controlling accessibility of the retroelements to the chromosome end. We have initiated a project to ask whether the telomere cap influences transposition to the chromosome end by asking whether mutations that modify the chromosome cap also influence the frequency of transposition. Currently stocks are being constructed that will be used in this assay. We are also trying to establish the technique of immunofluorescence-fluorescence in situ hybridization (IF-FISH) in the lab in order to identify chromatin structure in specific telomere regions and ask whether changes in these regions affect telomere maintenance. After irradiation, mutations in the mutator gene, mu2, increase the frequency of mutant chromosomes that have lost a telomere and reconstituted a structure that protects the chromosome end. MU2 accumulates at DNA repair foci, and mutations in the gene interfere with the formation of foci. The carboxy end of the MU2 protein interacts with the chromatin proteins HP1 and phosphorylated histone H2Av, the Drosophila orthologue of human H2AX, while the amino terminus interacts with the Mre11/Rad50/Nbs1 complex. Thus, MU2 appears to act as a scaffold for repair foci. This is consistent with sequence alignments, which suggest that mu2 is homologous to the human MDC1 gene. In an attempt to understand the interaction of factors that control telomere stability, we are making double mutation combinations of mu2 with a mutation that decreases telomere stability and increases telomere fusions. We will test both the generation of new telomeres and the frequency of telomere fusions in these double mutant lines.