1. Role of chromatin modifications in transcriptional regulation and repairing DNA breaks
In eukaryotic cells, chromosomes are packaged to specific protein-DNA complex called “chromatin”. Chromatin modifications and remodeling have been perceived as an integral part of daily chromosome transactions including transcriptional regulation and chromosomal damage recognition and repair processes. I have defined how transcription factor (HNF3) drives expression of liverspecific genes during hepatocyte differentiation in a murine system. In addition, my results established a new paradigm describing how a transcription factor modulates chromatin architecture conducive to transcription at specific gene promoters during cell differentiation (McPherson, 1993, Shim, 1998). I also demonstrated how the enigmatic protein kinase CDK9 controls elongation of RNA polymerase (Shim, 2002), but not the initiation of transcription, and thereby dictates expression of heat shock genes in C. elegans. Recently, I have devoted my research to defining how chromatin remodeling contributes to genome integrity. I showed that evolutionary conserved RSC chromatin remodeler is targeted to the DSB created in vivo and involved in repair of DNA double strand breaks by non-homologous end joining (NHEJ) and homologous recombination (HR) (Shim, 2005, 2007). I also found that RSC-dependent chromatin remodeling at DSB promotes the accessibility of Mre11 and Ku proteins to the DSBs, facilitating end resection and cohesin loading (Shim, 2007, Oum, 2011). I discovered that acetylated histone H3 impedes extensive repair synthesis (Che, 2015), broadening the effect of chromatin modification on post-invasion step of DNA recombination. Additionally, I identified mammalian XPF and XPF homologue have distinct roles in replicationcoupled and uncoupled inter-strand crosslink repair (Seol, 2018).
a. McPherson C.E., Shim E.Y., Friedman D.S. and Zaret K.S. (1993) An active tissue-specific enhancer and bound transcription factors existing in a precisely positioned nucleosomal array. Cell. 75: 385-398.
b. Shim E.Y., Woodcock C. and Zaret K.S. (1998) Nucleosome positioning by the winged helix transcription factor HNF3. Genes Dev. 12: 5-10.
c. Shim E.Y., Walker A. and Blackwell, T.K. (2002) CDK9/CyclinT (P-TEFb) is required in two postinitiation pathways for transcription in C. elegans embryo. Genes Dev. 16: 2135-2146.
d. Shim E.Y., Ma J.L., Oum J.-H., Yanez Y. and Lee S.E. (2005) The yeast chromatin remodeler RSC complex facilitates end joining repair of DNA double-strand breaks. Mol. Cell. Biol. 25: 3934-3944. PMCID: PMC1087737
e. Shim E.Y., Hong S.J., Oum J.H., Yanez Y., Zhang Y. and Lee S.E. (2007) RSC mobilizes nucleosomes to improve accessibility of repair machinery to the damaged chromatin. Mol. Cell Biol. 27: 1602-1613. PMCID: PMC1820475
f. Oum J. H, Seong C, Kwon Y, Ji J. H, Sid A, Ramakrishnan S, Ira G, Malkova A, Sung P, Lee S. E and Shim E.Y. (2011) RSC Facilitates Rad59-Dependent Homologous Recombination Between Sister Chromatids by Promoting Cohesin Loading at DNA Double-Strand Breaks. Mol. Cell. Biol. 39:3024-3039
g. Seol J. H., Holland C., Li X., Kim C., Li F., Medina-Rivera M., Elchmiller R., Gallardo I.F., Finkelstein I.J., Hasty P., Shim E.Y., Surtees J.A. and Lee S.E. (2018) Distinct roles of XPF-ERCC1 and Rad1-Rad10-Saw1 in replication-coupled and uncoupled inter-strand crosslink repair. Nat. Commun. 9:2025
h. Sohn J., Lee S.E., Shim E. Y. (2023) DNA Damage and Repair in Eye Diseases. Int J Mol Sci. 24: 3916
2. Mechanisms of DNA end resection and regulation of repair pathway choice
Chromosomal breaks, if not repaired correctly, fuels mutagenesis and chromosomal rearrangements. The key regulatory step in DNA damage repair and signaling is “DNA resection” that suppresses chromosomal instability upon exposure of environmental and endogenous genotoxins and various genetic diseases with cancer predisposition. Our past research endeavors have made several landmark discoveries about resection mechanism: (1) We have identified the key components of end resection and how these factors coordinate to achieve efficient end resection in yeast (Cell, 2008). (2) We have discovered that Mre11/Rad50/Xrs2 complex antagonizes Ku accumulation and thereby stimulates Exo1 nuclease activity (EMBO, 2012). (3) We have also identified the key components of end resection and how these factors coordinate to achieve efficient end resection in yeast (Cell, 2008 and NSMB, 2011). Most importantly, the resection genes and pathways we identified have been always validated for their roles in equivalent process in other model organisms including fission yeast, frogs, mouse and humans. We also demonstrated that the presence of MH sequence on flanking break sites dramatically increases the frequency of chromosomal translocations using an innovative genetic approach that induces a reciprocal translocation in real time by creating two site-specific DNA double strand breaks at separate chromosomes (PLOS genetics, 2012). Recently we showed that microhomology-mediated end joining induces hyper-mutagenesis at break junctions (PLOS genetics, 2017) and cellular assays of DNA repair pathways (Genes, 2019).
a. Shim, E.Y., Chung, W.H., Nicolette, M.I., Zhang,Y., Davis, M., Zhu, Z., Paull, T.T., Ira, G. and Lee, S.E. (2010) Saccharomyces cerevisiae Mre11/Rad50/Xrs2 and Ku proteins regulate association of Exo1 and Dna2 with DNA breaks. EMBO Journal. 29: 3370-3380
b. Chen, X., Niu, H., Chung W.H., Zhu, Z., Papusha, A., Shim, E.Y., Lee, S.E., Sung, P. and Ira, G. (2011) Cell cycle regulation of DNA double-strand break end resection by Cdk1-dependent Dna2 phosphorylation. Nat. Struct. Mol. Biol. 18: 1015-1019.
c. Sinha, S., Li, F., Villarreal, D., Shim, J.H., Yoon, S., Myung, K., Shim, E.Y. and Lee, S.E. (2017) Microhomology-Mediated End Joining induces hyper-mutagenesis at breakpoint junctions. PLOS genetics. 13(4):e1006714.
d. Lee, K.H., Ji, J.H., Yoon, K.H., Che, J., Seol, J, Lee, S.E. and Shim, E.Y. (2019) Microhomology selection for microhomology mediated end joining in Saccharomyces cerevisiae. Genes. 10: 284
e. Li, F., Wang, Q., Seol, J.H., Che, J., Lu, X., Shim, E.Y., Lee, S.E. and Niu, H. (2019) Apn2 resolves blocked 3’ ends and suppresses Top1-induced mutagenesis at genomic rNMP sites. Nat Struct Mol Biology. 26: 155-163.
3. Development of DNA Double Strand Break Dosimeter
Accurate calculation of radiation dose is essential for successful radiation therapy to cure cancers. However, the standard dosimeter estimates physical and chemical effects of radiation but it does not reliably measure biological effect of radiation such as cell death or DNA damage. The lack of dosimeter that faithfully reflects biological effect prompted us to develop a novel DNA dosimeter detecting the key lesions induced by radiation, DNA double strand breaks (DSBs) (Medical Physic, 2018). The current study validates the ability of a DNA DSB dosimeter to successfully reflect Relative Biological Effects (RBE) of low and high energy x-ray radiation and correlates with cell survivals and in vivo DNA double strand breaks formation by neutral comet assay. Results support the benefit of DSB dosimeter to detect radiation dosage relevant to RBE and to underscore the effect of radiation energy on the extent of DNA DSBs (Red Journal, in preparation)
a. Obeidat M, McConnell KA, Li X, Bui B, Stathakis S, Papanikolaou N, Rasmussen K, Ha CS, Lee SE, Shim EY*, Kirby N*. (2018) DNA double-strand breaks as a method of radiation measurements for therapeutic beams. Medical Physics. 45: 3460-3465.
b. Li X, McConnell KA, Che J, Ha CS, Lee SE, Kirby N and Shim EY. (2020) DNA dosimeter Measurement of Relative Biological Effectiveness for 160 kVp and 6 MV X-rays. Radiation Research. 194: 173-179.
