This International Collaboration in Chemistry (ICC) award in the Chemistry of Life Processes (CLP) program in the Division of Chemistry, with supplemental funds from NSF's Office of International Science and Engineering (OISE), supports work by Professor Hanbin Mao at Kent State University and Professor Hiroshi Sugiyama of Kyoto University, Japan, who is funded by the Japan Society for the Promotion of Science (JSPS), to carry out fundamental studies on the identification and characterization of novel structures in the human telomere regions. Located at the end of chromosomes, the telomere plays a critical role in cancers and senescence via DNA quadruplex structures that contain four DNA strands. Identification of novel structures other than quadruplexes is instrumental in understanding the fundamental processes of senescence and genesis of cancer. Single molecular techniques, such as laser tweezers and atomic force microscopy will be used to reveal the presence of the novel structures, to investigate the interaction of these structures with ligands, and to evaluate the effect of these structures on telomerase, an enzyme that maintains the telomere length. These methods will be complemented by molecular simulation and conventional bulk assays such as native gel electrophoresis, thermal denaturation, circular dichroism, and surface plasma resonance (SPR). The results from this three-year project, the broader impacts of which include the training of undergraduate and graduate students, can help to identify new therapeutic targets and agents to fight against diseases such as cancer.
Project Report 2010-2014 Hanbin Mao (a) NSF award number: CHE-1026532; Amount: $278,000; Period of support: 10/01/2010-9/30/2013. With no-cost extension to 9/30/2014. (b) Title of the project: International Collaboration in Chemistry: Investigation of Novel Telomeric Structures and Their Effects on Human Telomerase. (c) Summary of the results of the completed project: This international collaboration in chemistry (ICC) award supports research by Prof. Hanbin Mao (US) and Prof. Hiroshi Sugiyama (Japan) to identify and characterize novel structures in the human telomere region. Located at the end of chromosomes, telomeres can host DNA G-quadruplex structures where G-quartets consisting of four Guanine residues linked by Hoogsteen base pairs are stacked into a complex structure. Telomeric G-quadruplexes have been proposed to play important roles leading to the senescence and genesis of cancers. This NSF grant aims to identify novel structures, other than regular G-quadruplexes, that could also be involved in the senescence and cancer genesis. To do this, highly sensitive single molecular techniques, such as laser tweezers and atomic force microscopy, have been proposed to reveal the presence of the novel structures, to investigate the interaction of these structures with ligands, and to evaluate the effect of these structures on telomerase, an enzyme that maintains the telomere length. During this grant period (2010-2014), we have made the following accomplishments. We observed the presence of a new species that is consistent with a G triplex conformation in the human telomeric region. We investigated the binding interaction between a G quadruplex and a ligand in a new laser tweezers platform in which a force jump method was introduced. We designed a click chemistry approach to investigate the folding and unfolding trajectories of telomeric G-quadruplex under different folding or unfolding trajectories. We measured multi-dimensional distances in a G-quadruplex species to reveal specific 3D conformation of the structure at the single molecular level. We identified different species formed in long DNA telomeric sequences and measured transition kinetics between these species. We found the nascent RNA produced by transcription process can facilitate the formation of DNA G-quadruplex. During this period, we have published 13 peer-reviewed papers as listed below, 1. Journal of the American Chemical Society, 2014, 136, 13967-13970. 2. Nucleic Acids Research, 2014. DOI:10.1093/nar/gku416. 3. Nucleic Acids Research, 2014, 42, 5755-5764. 4. ChemBioChem, 2013, 14, 1931-1935. 5. Reviews in Analytical Chemistry, 2013, 32, 197-208. 6. Journal of the American Chemical Society, 2013, 135, 2235−2241. 7. Nucleic Acids Research, 2013, 41, 3915-3923. 8. Chemical Record, 2013, 13, 102–116. 9. Journal of the American Chemical Society, 2012, 134, 12338–12341. 10. Analytical Chemistry, 84, 5298-5303, 2012. 11. Chemical Communications, 2012, 48, 2006-2008. 12. Journal of the American Chemical Society, 2012, 134, 5157-5164. 13. Nature Chemistry, 2011, 3, 782-787. In addition, this NSF grant has allowed us to train five high school students, Ashley Paydock (Ravenna high school, Ravenna, OH), Mira Scarnecchia (Hudson high school, Hudson, OH), Jill Mao (Fairfax high school, Fairfax, VA), Danylo Lavrentovich (Hudson high school, Hudson, OH), Jia Wei (Hudson high school, Hudson, OH); four college students, Brittany Pleat (KSU); Nathan Holliday (Georgetown College); Athena Guo (KSU); William J Maximuck (Millersville University); and five graduate students, Deepak Koirala, Zhongbo Yu, Soma Dhakal, Philip Yangyuoru, and Chiran Ghimire at Kent State University. As a part of international collaboration, these students interact frequently with professor Hiroshi Sugiyma and his graduate students, Yuta Sannohe and Seigi Yamamoto in Japan. Soma attended the third international conference on G-quadruplexes at Italy in 2011. Deepak spent one month in Sugiyamaâ€™s lab, learning AFM imaging on DNA structures. Deepak attended Gordon conference 2013 where he met G-quadruplex researchers, Naoki Sugimoto and Cynthia Burrows. Such an invaluable international experience is beneficial to instill young scientists a global perspective to a research effort, which is highly important for current trend of globalization. In addition, the involvement of undergraduate students in the research project certainly will allure them into the field of science, which US desperately needs to strengthen its competitiveness.