Most eukaryotic chromosome ends terminate in structures of repetitive DNA sequences and associated proteins, called telomeres. Telomeres allow cells to distinguish natural chromosome ends from damaged DNA and to protect chromosomes against degradation and fusion. Telomere integrity in cells thus plays an essential role in controlling genomic stability. Several key mechanisms are involved in telomere maintenance, including telomerase, telomere associated proteins and key DNA repair proteins. Loss of telomere protection at chromosome ends is frequently observed in elder populations, cellular senescence, and premature aging syndromes. Furthermore, telomere dysfunction contributes to genomic instability that leads to cell death, cell proliferation defects, and malignant transformation, which might in turn contribute to age related-disorders and a higher incidence of cancer during aging.? ? Replicative senescence and oxidative stress have been implicated in aging. It has been shown that oxidative stress affects telomere dependent replicative lifespan of primary human cells in culture. Our lab is currently examining the role of oxidative damage and its corresponding repair mechanisms at telomere. The paradigm used in the studies is based on three levels of measure: (1) Dissecting DNA repair pathways, e.g. base excision repair (BER) that contribute to repair of oxidative damage at telomeres; (2) Investigating the deteriorating effect of oxidative damage on telomere integrity and genome stability; (3) Exploring the environmental factors that cause oxidative stress at telomeric DNA that influences telomere maintenance at cellular and organism levels, especially their impacts on several sensitive human tissues. The overall effort will facilitate our understanding in how oxidative stress may impact telomere integrity and thus aging and age-related disease. ? ? For this fiscal year, our primary effort has been to (1) Set up a functional laboratory, including purchasing lab equipment, recruiting technician and postdoctoral fellows, training lab staff to perform laboratory experiments, keep records of experimental procedures, and conduct laboratory management; (2) Elucidate the role of oxidative damage and its repair at telomeres. We mainly focus on the characterization of BER pathways, i.e. DNA glycosylase(s) in excising damaged telomeric bases caused by Reactive Oxygen Species (ROS) in vivo. We are in the process to identify if oxidized base modification/damage lead to telomere dysfunction in murine and human cells and if DNA glycosylase(s) are critical in repair oxidized base modification/damage. We also try to establish several methodologies that can be used to identify special types of base modifications/damage at telomeres. The latter effort was carried out through the collaboration with Drs. Vilhelm Bohrs and David Wilsons laboratories. To study the environmental factors that may result in oxidative stress at telomeric DNA and thus influence telomere maintenance, we are investigating if smoking causes oxidative base modification/damage at telomeres that leads to telomere shortening or dysfunction in human lung cell lines; (3) Investigate the role of a DNA repair protein, Fanconi C, in telomere length regulation and bone marrow failure in human Fanconi anemia. We have initiated the analysis of telomere length and telomere damage in Fanconi C deficient murine bone marrow cells; (4) Establish internal and external collaborations: The characterization of DNA repair pathways and telomere associated proteins in telomere maintenance is through collaboration with several groups, including Drs.Vilhelm Bohr, Patricia J. Gearhart, Michael Seidman, David M. Wilson, III, and Robert M. Brosh (LMG) and Drs. David Gilley and Laura Hanelline (Indiana University), Yisong Wang and Hays McDonald (Oak Ridge National Laboratory); (5) Present our work in the internal and international conferences: We reported our work in the Telomere and Telomerase International Conference, New York and annual NIA retreat; (6) Establish multi-color Fluorescent In Situ Hybridization (mFISH) technique at LMG. This technique allows the detection of gross chromosome abnormalities, e.g. aneuploidy, chromosomal translocation, and large deletion or insertion.
