https://scholarworks.unist.ac.kr/handle/201301/5 Wed, 08 Apr 2026 00:40:33 GMT 2026-04-08T00:40:33Z https://scholarworks.unist.ac.kr/handle/201301/90890 Title: Xenopus rsph6a.L: A Conserved Model for Human RSPH4A associated Ciliopathy Author(s): Hwang, Jae Hyung Abstract: Mutations in the radial spoke head protein RSPH4A are a primary cause of Primary Ciliary Dyskinesia (PCD). While mouse models serve as the gold standard for confirming pathogenicity, high costs and invasive procedures limit their utility for initial high-throughput screening. Conversely, while foundational studies in Chlamydomonas reinhardtii have provided structural insights, their evolutionary distance from mammals limits direct translational application. To address these limitations, we evaluated Xenopus laevis as a cost-effective vertebrate model that offers distinct experimental advantages. Although Xenopus possesses a single rsph6a.L gene, our phylogenetic and structural analyses reveal that it shares high conservation with mammalian RSPH4A and RSPH6A, retaining functional domains commonly affected in human patients. We depleted rsph6a.L using CRISPR-Cas12a and Morpholino oligonucleotides. Immunofluorescence analysis revealed that ciliary axoneme assembly remained intact; however, bead flow assays demonstrated a significant reduction in fluid flow velocity. This defect was attributed to a loss of planar beating directionality and a transition to aberrant rotational motion. These findings demonstrate that rsph6a.L is essential for the coordination of mucociliary clearance and validate the Xenopus system as a conserved, rapid screening tool for assessing RSPH4A-associated PCD pathogenicity. Major: Department of Biological Sciences Sat, 31 Jan 2026 15:00:00 GMT https://scholarworks.unist.ac.kr/handle/201301/90890 2026-01-31T15:00:00Z https://scholarworks.unist.ac.kr/handle/201301/90887 Title: CXXC5 deletion enhances tumorigenesis by impairing DNA damage responses in CRC Author(s): Oh, Seungmin Abstract: CXXC5 (RINF) is a CpG-binding epigenetic regulator implicated in transcriptional control and cellular stress responses; however, its role in epithelial DNA damage repair and colorectal tumorigenesis remains incompletely understood. Because maintenance of genomic stability in the intestinal epithelium relies heavily on efficient double-strand break (DSB) repair, we investigated whether CXXC5 contributes to non-homologous end joining (NHEJ) and influences APC-driven colorectal tumor development.Analysis of publicly available colorectal cancer datasets revealed that CXXC5 expression is elevated in tumor tissues compared with normal counterparts. Consistent with this observation, APCmin/+ mice exhibited increased Cxxc5 protein levels in tumor regions relative to adjacent non-tumor intestine. To examine the epithelial-specific function of CXXC5 in vivo, we generated intestinal epithelium–specific Cxxc5 knockout mice using Villin-Cre, with or without the APCmin/+ background, and confirmed efficient depletion of CXXC5 in the intestinal epithelium by Western blotting and immunohistochemistry.Functionally, epithelial loss of Cxxc5 was associated with earlier clinical decline and increased colorectal tumor burden in APCmin/+ mice. These changes were accompanied by increased colon weight without additional shortening of colon length, as well as enhanced tumor multiplicity, larger tumor size, and more severe histopathological abnormalities. To explore a potential link to DNA repair, chromatin fractionation assays performed in SW620 colorectal cancer cells showed reduced chromatin recruitment of Ku70 and DNA-PKcs, along with sustained γH2AX accumulation following DNA damage, suggesting impaired resolution of DSBs via the NHEJ pathway.Together, these findings indicate that CXXC5 contributes to DNA damage responses in intestinal epithelial cells and that its loss is associated with defective recruitment of specific NHEJ factors and enhanced APC-driven colorectal tumor development. This study suggests a role for CXXC5 in epithelial genome maintenance under conditions of chronic genomic stress. Major: Department of Biological Sciences Sat, 31 Jan 2026 15:00:00 GMT https://scholarworks.unist.ac.kr/handle/201301/90887 2026-01-31T15:00:00Z https://scholarworks.unist.ac.kr/handle/201301/90888 Title: Transcriptional factors PU.1 and CEBP/α drive direct reprogramming fibroblasts into tissue resident macrophages Author(s): Lee, Joo Chan Abstract: Abstract Macrophages are highly plastic innate immune cells whose identities are established by lineage-determining transcription factors and tissue-specific enhancer landscapes. The ETS family transcription factor PU.1 and the CCAAT/enhancer-binding protein C/EBPα are core regulators of myeloid differentiation and macrophage fate. Previous studies have shown that enforced expression of these factors can directly convert fibroblasts and lymphoid progenitors into macrophage-like cells. However, the epigenomic properties of fibroblast-derived macrophage-like cells, particularly their relationship to tissue-resident macrophage (TRM) enhancer programs, remain incompletely understood. In this study, we established a doxycycline-inducible (Tet-On) PU.1/C-EBPα overexpression system in NIH-3T3 fibroblasts and characterized their direct conversion into macrophage-like cells. Induction of PU.1 and C/EBPα led to marked morphological remodeling, acquisition of macrophage surface markers CD11b and F4/80, and inflammatory responsiveness to lipopolysaccharide stimulation, as demonstrated by increased TNF-α and IL-1β expression. Furthermore, by integrating enhancer-centered chromatin profiling strategies with published TRM datasets, reprogrammed fibroblasts were shown to acquire macrophage-associated and TRM-like enhancer features. Super-enhancer mapping identified Hivep3 as a candidate transcription factor linked to TRM regulatory programs. Collectively, these findings demonstrate that inducible PU.1/C-EBPα expression is sufficient to generate functional macrophage-like cells from fibroblasts and partially recapitulate TRM enhancer architecture. This platform provides a controllable experimental system for dissecting macrophage gene regulatory networks and for identifying transcriptional regulators of tissue-resident macrophage identity. Major: Department of Biological Sciences Sat, 31 Jan 2026 15:00:00 GMT https://scholarworks.unist.ac.kr/handle/201301/90888 2026-01-31T15:00:00Z https://scholarworks.unist.ac.kr/handle/201301/90889 Title: Oxygen-Dependent Phase Separation of PHD3 Links the Hypoxic Microenvironment to Mitochondrial Metabolic Reprogramming in ccRCC Author(s): Lim, Soyeon Abstract: Clear cell renal cell carcinoma (ccRCC) is characterized by profound metabolic dysregulation, with both PHD3 and pyruvate carboxylase (PC) independently implicated in disease progression. While each influences patient outcomes, a direct mechanistic interplay between these two regulators has remained elusive. Here, we uncover a novel regulatory axis involving PHD3 and PC by identifying an unexpected subcellular behavior of PHD3, namely its dual localization to the cytosol and the mitochondrial matrix. We show that mitochondrial import of PHD3 is driven by liquid–liquid phase separation (LLPS), a biophysical process modulated by PHD3 hydroxylase activity and oxygen levels. Once in the matrix, PHD3 directly hydroxylates PC, suppressing its enzymatic activity. In ccRCCs with elevated PHD3 expression, this modification restricts anaplerotic flux into the tricarboxylic acid (TCA) cycle, leading to impaired proliferation, reduced metastasis, and enhanced apoptosis. Together, our findings provide a new framework for targeting cancer metabolism by establishing a previously unrecognized mechanistic link between PHD3- mediated oxygen sensing within the tumor microenvironment (TME) and the regulation of ccRCC mitochondrial metabolism through LLPS-driven translocation. Major: Department of Biological Sciences Sat, 31 Jan 2026 15:00:00 GMT https://scholarworks.unist.ac.kr/handle/201301/90889 2026-01-31T15:00:00Z