Genomic instability is now known to be an early step in cancer and can generate many of the chromosome rearrangements leading to tumor cell progression. We have demonstrated that even tumor cells that express telomerase can have relatively high rates of telomere loss, and that this spontaneous telomere loss is an important mechanism for chromosome instability in cancer. We have also demonstrated that telomeres that have been lost in mammalian cells can be restored through a process called chromosome healing, and that chromosome healing can prevent chromosome instability. Understanding the factors and genes that influence telomere loss and chromosome healing is therefore important for understanding cancer and may provide new approaches for cancer therapy.
Major Goals: (1) understand the mechanisms of genomic instability and their role in human disease; (2) investigate the role of chromosome healing in preventing chromosome instability; (3) determine the relationship between subtelomeric chromatin structure, gene silencing, and telomere function; (4) establish new approaches for cancer therapy by manipulating the regulation of chromosome healing.
Chromosome instability resulting from spontaneous telomere loss in human cancer. Despite the expression of telomerase, we have found that human cancer cells have a high rate of spontaneous telomere loss, which can generate many of the types of chromosome rearrangements commonly found in cancer cells. We are now actively involved in the study of the mechanisms and genes responsible for this high rate of spontaneous telomere loss and its role in cancer.
Chromosome instability resulting from double-strand breaks (DSBs) near telomeres in mouse embryonic stem (ES) cells and human cancer cells. Ionizing radiation can induce chromosome instability through the formation of DNA double-strand breaks (DSBs). Our results show that a single DSB near a telomere causes telomere loss and initiates chromosome instability in mouse ES cells and human tumor cells. Our studies with human tumor cells show that this telomere loss results from deficient DSB repair in regions near telomeres, which causes fusion of sister chromatids leading to chromosome instability. We are currently studying the mechanisms and genes responsible for this decreased repair capability near telomeres and the resulting complex chromosome rearrangements that result.
Restoration of lost telomeres and its role in preventing chromosome instability. We have demonstrated that both mouse ES cells and human tumor cells can restore lost telomeres through a process called chromosome healing. We have also demonstrated that chromosome healing in mouse ES cell lines can occur through both telomerase-dependent and independent pathways, and that it can prevent chromosome instability due to telomere loss. We are currently investigating the role of Pif1 in the regulation of chromosome healing, since yeast Pif1 inhibits chromosome healing, and prevents it from interfering with DSB repair. Understanding the regulation of chromosome healing is important, since upregulation of chromosome healing could specifically sensitize cancer cells to ionizing radiation and limit the chromosome instability associated with development of resistance of cancer cells to chemotherapeutic agents.
Mechanisms of silencing of genes located near telomeres. Using our mouse ES cells and transgenic mice containing telomeric transgenes, we have demonstrated the silencing of genes located near telomeres, termed telomere position effect (TPE). TPE has been proposed to be involved in cancer and aging, so we are using this system to identify the mechanisms and genes involved. We have also used this system to screen an shRNA library for genes that can inhibit silencing, through selection for re-expression of silenced selectable marker genes. These studies have identified a large number of chromatin modifications and proteins involved in subtelomeric chromatins structure, many of which are also influence telomere function and DNA repair.
Fouladi, B, Miller, D., Sabatier, L. and Murnane, J.P. The role of spontaneous telomere loss in chromosome instability in a human tumor cell line. Neoplasia 2:540-554 (2000).
Lo, A. W. I., Sprung, C., Fouladi, B., Pedram, M., Sabatier, L., Ricoul, M., Reynolds, G. E. and Murnane, J. P. Chromosome instability as a result of double-strand breaks near telomeres in mouse embryonic stem cells. Mol. Cell. Biol. 22:4836-4850 (2002).
Lo, A. W. I., Sabatier, L., Fouladi, B., Pottier, G, Ricoul, M., and Murnane, J. P. DNA amplification by breakage/fusion/bridge cycles initiated by spontaneous telomere loss in a human cancer cell line. Neoplasia 4:531-538 (2002).
Bai, Y. and Murnane, J. P. Telomere instability in a human tumor cell line expressing a dominant-negative WRN protein. Human Genetics, 113:337-347 (2003).
Bai, Y. and Murnane, J. P. Telomere instability in a human tumor cell line expressing a NBS1 with mutations at sites of phosphorylation by ATM. Mol. Can. Res. 1:1058-1069 (2003).
Murnane, J. P. and Sabatier, L. The role of telomeres in chromosome instability and cancer. BioEssays, 26:1164-1174 (2004).
Sabatier, L., Ricoul, M., Pottier, G., Mathieu, N. and Murnane, J. P. The loss of a single telomere can result in instability of multiple chromosomes in a human tumor cell line. Mol. Can. Res., 3:139-150 (2005).
Pedram, M., Sprung, C. N., Gao, Q., Lo, A. W. I., Reynolds, G. E., and Murnane, J. P. Telomere position effect and silencing of transgenes near telomeres in mouse. Mol. Cell. Biol., 26:1865-1878 (2006).
Bailey, S. M. and Murnane, J. P. Telomeres, Chromosome instability and Cancer. Nucleic Acids Res., Nucleic Acids Res., 34: 2408-2417 (2006).
Murnane, J. P. Telomeres and chromosome instability. DNA repair, 5:1083-1092 (2006).
Gao, Q, Reynolds, G. E., Innes, L., Pedram, M., Jones, E., Junabi, M., Gao, D.-W., Ricoul, M., Sabatier, L., Van Brocklin, H., Franc, B. L., and Murnane, J. P. Telomeric transgenes are silenced in adult mouse tissues and embryo fibroblasts, but are expressed in embryonic stem cells . Stem Cells, 25:3085-3092 (2007).
Ho, C. Y. S., Murnane, J. P., Yeung, A. K. Y., Ng, H. K., and Lo, A. W. I. Telomeres acquire distinct heterochromatin characteristics during siRNA-induced RNA interference in mouse cells. Current Biol., 18:183-187 (2008).
Gao, Q, Reynolds, G. E., Wilcox, A., Miller, D., Cheung, P., Artandi, S., and Murnane, J. P. Telomerase-dependent and independent mechanisms of chromosome healing in mouse embryonic stem cells. DNA Repair, 7:1233-1249 (2008).