Werner's syndrome (WS) is a homozygous recessive disease characterized by early onset of many characteristics of normal aging, such as wrinkling of the skin, graying of the hair, cataracts, diabetes, and osteoporosis. Because of the acceleration of aging in WS, the study of this disease will hopefully shed light on the degenerative processes that occur in normal aging. Cells from WS patients grow more slowly and senescence at an earlier population doubling than age-matched normal cells, possibly because these cells appear to lose the telomeric ends of their chromosomes at an accelerated rate. In general, WS cells have a high level of genomic instability, with increased amounts of DNA deletions, insertions, and rearrangements. These effects could potentially be the result of defects in DNA repair, replication, and/or recombination, although the actual biochemical defect remains unknown. We are comparing WRN to the other RecQ helicases. There are five human RecQ proteins and all are involved in the maintenance of genome stability. We are especially interested in defining the unique and shared roles of WRN and the other RecQ helicases in double strand break repair. Confocal microscopy is used as a means to investigate the dynamic behavior of WRN and its interacting partners. There are multiple pathways whereby a cell can repair a double strand break (DSB) and recently we investigated WRN's role in DSB repair pathway choice (Homologous Recombination (HR) vs classical Non-Homologous End Joining (c-NHEJ) vs Alternative NHEJ (alt-NHEJ)). We find that WRN regulates resection at the 5' ends of DSBs, and influences the choice between c-NHEJ and alt-NHEJ. In mouse cells lacking Wrn and Trf2, we found that alt-NHEJ is favored leading to an increased frequency of telomere fusions. Thus, alt-NHEJ may be driving genomic instability in WRN-deficient cells. The implications from this work are that small molecule inhibitors of alt-NHEJ may have therapeutic benefit in WRN-deficient cells by preventing genomic rearrangements and genome stability. We have also explored the role of WRN expression in breast cancer. Breast cancer is the leading cause of cancer deaths in women aged 20-59 worldwide, and the second leading cause of death for women in the US. Breast cancer predisposition is polygenic and influenced by environmental factors. There are both familial and sporadic forms of breast cancer, and while significant progress has been made identifying high (BRCA1) and low risk breast cancer-associated genes, there are still many genes yet to be identified. Recent studies identified BLM and RECQL1 as potential breast cancer susceptibility loci. Thus, we are currently investigating the roles that RecQ proteins may play in breast cancer and in response to chemotherapy. In general, cells with defects in DNA repair are at increased risk of transformation. Recently, in collaboration with Dr. Madhusudan (Nottingham, UK), we identified elevated expression of BLM, RECQL4 and RECQL5 mRNA, and low expression of WRN and RECQL1 mRNA in patients with aggressive breast tumors and poor breast cancer specific survival. Aggressive tumors also displayed aberrant and/or extranuclear immunostaining of one or more RecQ helicase proteins. We also studied the effects of camptothecin (a Topoisomerase I (TopoI) poison) on the stability and expression of RecQ proteins and TopoI in breast cancer cell lines. Interestingly, WRN was degraded, but TopoI was not, and this correlated with camptothecin sensitivity. There are very few proteins which predict clinical sensitivity to drugs; however WRN protein stability appears to be one. To extend these results, we will examine WRN and its protein networks in breast cancer cell lines and in response to other Topo1 inhibitors. This and other experimental approaches will be used to elucidate in molecular detail the mechanism by which RecQ proteins contribute to or modulate breast cancer.
| Maity, Jyotirindra; Das, Biswadip; Bohr, Vilhelm A et al. (2018) Acidic domain of WRNp is critical for autophagy and up-regulates age associated proteins. DNA Repair (Amst) 68:1-11 |
| Lee, Mina; Shin, Soochul; Uhm, Heesoo et al. (2018) Multiple RPAs make WRN syndrome protein a superhelicase. Nucleic Acids Res 46:4689-4698 |
| Shamanna, Raghavendra A; Croteau, Deborah L; Lee, Jong-Hyuk et al. (2017) Recent Advances in Understanding Werner Syndrome. F1000Res 6:1779 |
| Shin, Soochul; Lee, Jinwoo; Yoo, Sangwoon et al. (2016) Active Control of Repetitive Structural Transitions between Replication Forks and Holliday Junctions by Werner Syndrome Helicase. Structure 24:1292-1300 |
| Tippana, Ramreddy; Hwang, Helen; Opresko, Patricia L et al. (2016) Single-molecule imaging reveals a common mechanism shared by G-quadruplex-resolving helicases. Proc Natl Acad Sci U S A 113:8448-53 |
| Shamanna, Raghavendra A; Lu, Huiming; Croteau, Deborah L et al. (2016) Camptothecin targets WRN protein: mechanism and relevance in clinical breast cancer. Oncotarget 7:13269-84 |
| Khadka, Prabhat; Croteau, Deborah L; Bohr, Vilhelm A (2016) RECQL5 has unique strand annealing properties relative to the other human RecQ helicase proteins. DNA Repair (Amst) 37:53-66 |
| Khadka, Prabhat; Hsu, Joseph K; Veith, Sebastian et al. (2015) Differential and Concordant Roles for Poly(ADP-Ribose) Polymerase 1 and Poly(ADP-Ribose) in Regulating WRN and RECQL5 Activities. Mol Cell Biol 35:3974-89 |
| Edwards, Deanna N; Machwe, Amrita; Chen, Li et al. (2015) The DNA structure and sequence preferences of WRN underlie its function in telomeric recombination events. Nat Commun 6:8331 |
| Croteau, Deborah L; Popuri, Venkateswarlu; Opresko, Patricia L et al. (2014) Human RecQ helicases in DNA repair, recombination, and replication. Annu Rev Biochem 83:519-52 |
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