https://scholar.dgist.ac.kr/handle/20.500.11750/11738 2026-08-07T21:42:17Z 2026-08-07T21:42:17Z 강은호 Ashim Janbolat Ji, Sangho Han, Min Ae Yu, Wookyung Kim, Sung Jae Hong, Eunmi Moon, Cheil Lee, Chang-Hun https://scholar.dgist.ac.kr/handle/20.500.11750/60516 2026-07-27T01:10:17Z 2026-06-30T15:00:00Z

Title: Expression and purification of the ectodomain of erythropoietin receptor fused to mCitrine or mTFP1 fluorescent protein in Escherichia coli Author(s): 강은호; Ashim Janbolat; Ji, Sangho; Han, Min Ae; Yu, Wookyung; Kim, Sung Jae; Hong, Eunmi; Moon, Cheil; Lee, Chang-Hun Abstract: The erythropoietin receptor (EPOR) is a single-pass transmembrane protein that homo-dimerizes upon binding with its renal ligand erythropoietin (EPO) to trigger downstream signaling. Its extracellular ectodomain mediates ligand binding. Therefore, a fusion protein of the EPOR ectodomain can be useful for various in vitro assays, such as a binding assay with an EPO-like peptide, if overexpressed in Escherichia coli (E.coli). In this study, we hypothesized that fusion proteins of the EPOR ectodomain with mCitrine or mTFP1, expressed in bacteria, could enable in vitro Fo & uml;rster resonance energy transfer experiments. Two fusion proteins, EPOR-mCitrine and EPORmTFP1, were overexpressed in E. coli but obtained as inclusion bodies. Urea solubilization and stepwise dialysis yielded soluble fusion proteins. Circular dichroism spectroscopy revealed that EPOR-mCitrine had greater secondary structure content than EPOR-mTFP1. When combined with recombinant human EPO, the hydrodynamic radius of EPOR-mCitrine changed, as measured using dynamic light scattering, confirming binding. This interaction was further validated using isothermal titration calorimetry. We propose that bacterially produced EPOR-mCitrine is a useful in vitro tool for measuring EPO binding.

2026-06-30T15:00:00Z Pineda, Christopher Cohen, Erez Su, Beau Yeom, Jiwoo Kim, Jewoo Lee, Chang-Hun Coulombe, Pierre A. https://scholar.dgist.ac.kr/handle/20.500.11750/60362 2026-05-22T06:40:12Z 2026-04-30T15:00:00Z

Title: A role for Keratin 17 in Rac1-mediated DNA damage response in keratinocytes Author(s): Pineda, Christopher; Cohen, Erez; Su, Beau; Yeom, Jiwoo; Kim, Jewoo; Lee, Chang-Hun; Coulombe, Pierre A. Abstract: Keratin 17 (K17) is a stress-responsive intermediate filament protein that is upregulated in chronic skin diseases and in several carcinomas. We previously showed that K17 is induced in epidermal keratinocytes following exposure to DNA-damaging agents, promoting keratinocyte survival and chemically induced papilloma formation in mouse skin. Molecularly, K17 is recruited to the nucleus, where it impacts nuclear architecture, gene expression, and the DNA damage response (DDR). Here, we report on efforts to delineate K17-dependent processes during DDR by focusing on its interacting partners. Using mass spectrometry, we identified a network of K17-interacting Rho GTPase signaling proteins, including Rac1 and its activator Dock7. Biochemically, we confirmed that Rac1 and K17 interact directly in vitro and in A431 tumor keratinocytes, both at baseline and after ionizing radiation. We show that KRT17 deletion leads to decreased levels of Rac1 protein, its DNA damage-related effector TOP2A, and a Rac1-dependent decrease in cellular proliferation following ionizing radiation. Remarkably, key K17-dependent readouts are rescued by expression of constitutively active, but not dominant-negative, Rac mutants in KRT17 null A431 keratinocytes. These findings uncover a K17-Rac1-TOP2A signaling axis that promotes DDR and associated proliferation, with implications for cancer and chronic skin diseases.

2026-04-30T15:00:00Z Kwack, Mi Hee Kang, Eunho Kim, Jewoo Ji, Youngheum Ju, Hyeonchang Lee, Chang-Hun Sung, Young Kwan Kim, So Yeon Moon, Cheil https://scholar.dgist.ac.kr/handle/20.500.11750/60353 2026-05-11T09:10:14Z 2026-02-28T15:00:00Z

Title: MLPH-mediated activation of dermal papilla IGF-1 signaling drives human hair shaft elongation and anagen induction Author(s): Kwack, Mi Hee; Kang, Eunho; Kim, Jewoo; Ji, Youngheum; Ju, Hyeonchang; Lee, Chang-Hun; Sung, Young Kwan; Kim, So Yeon; Moon, Cheil Abstract: Introduction Hair loss (alopecia) is a multifactorial disorder that often causes distress. Approved therapies such as minoxidil and finasteride act indirectly and do not specifically target hair follicle (HF) cells. Erythropoietin (EPO), however, has been shown to activate dermal papilla (DP) cells via the erythropoietin receptor (EPOR), suggesting a potential role in hair follicle regeneration and hair growth. Objectives This study aimed to develop and validate Helix C-1–based EPO-derived peptides that activate DP cells and increase IGF-1 expression, while not inducing overt systemic erythropoietic effects (e.g., increases in red blood cell counts, reticulocytes, hemoglobin, or hematocrit) under the tested experimental conditions. Methods Peptides derived from the Helix C-1 region of EPO were synthesized and characterized by EPOR-binding affinity, CD spectroscopy, and ERK/AKT activation. In vitro, DP-cell metabolic activity, proliferation, and IGF-1 secretion were assessed. Ex vivo efficacy was evaluated by hair shaft elongation in hair follicle organ culture, and in vivo efficacy was tested in a murine depilation-induced anagen model with concurrent hematologic assessment to exclude erythropoiesis-related effects. Results The peptides increased DP-cell metabolic activity and proliferation, reduced oxidative stress, and enhanced IGF-1 production via EPOR-mediated ERK/AKT activation. They promoted hair shaft elongation ex vivo and promoted anagen entry in mice without significant changes in standard hematologic parameters under the tested dosing regimen. Conclusion These findings support the conclusion that MLPH promotes hair growth via an EPOR-linked, IGF-1–dependent mechanism in DP cells. Future pharmacokinetic and disease-model studies are warranted to evaluate its translational potential. © 2026 The Authors.

2026-02-28T15:00:00Z Cho, Jung-Ah Jeon, Sang Seo Choi, Go Woon Lee, Chang-Hun Kim, Sung-Jae https://scholar.dgist.ac.kr/handle/20.500.11750/60228 2026-04-15T18:01:47Z 2025-12-31T15:00:00Z

Title: Investigation on physical and physiological properties of extracellular vesicles derived from Enterococcus faecalis Author(s): Cho, Jung-Ah; Jeon, Sang Seo; Choi, Go Woon; Lee, Chang-Hun; Kim, Sung-Jae Abstract: Extracellular membrane vesicles (EVs) are nanosized particles that contain various molecules originating from their parental cells and are produced by all three domains of life, including bacteria. Bacterial EVs are known to contribute to bacterial infections and immune responses in various human diseases. Enterococcus faecalis is an opportunistic pathogen. In this study, we examined the physical and physiological properties of EVs generated by E. faecalis, including particle size, protein composition, and cytokine-inducing profiles. To this end, we isolated EVs from bacteria under different preparation processes, and also a certain condition with the addition of EGCG. First, the bacterial culture supernatants were directly ultracentrifuged (named "Rough"), or filtered through 0.45- or 0.22 mu m pore-sized membrane filters (named as "0.45 mu m" or "0.22 mu m," respectively). EVs from EGCG-treated bacteria were prepared using a 0.45 mu m pore-sized membrane filter and named "EGCG + 0.45 mu m." Each EV sample was subjected to DLS, SDS-PAGE, and cytokine array analyses. DLS results showed that the differently prepared EVs had distinct size distributions depending on the filtration process. SDS-PAGE results revealed unique protein profiles that differentiated EVs under each condition. Treatment of macrophages with each EV sample markedly increased cell viability and size. The cytokine profiles produced by macrophages in response to each EV preparation revealed both common and distinguishable factors. This study has significance in revealing aspects of the biological characteristics of EVs produced by E. faecalis, which have previously been largely unknown.

2025-12-31T15:00:00Z