Biology subjects

Kim, H. G.

Publications and source records attributed to Kim, H. G..

7 recordsLinked to original sources

bioRxiv2025

Chromatin Remodeling and Transcriptional Silencing Define the Dynamic Innate Immune Response of Tissue Resident Macrophages After Burn Injury.

Severe burn injury induces long-lasting immune dysfunction, but the molecular mechanisms underlying this phenomenon remain unclear. We hypothesized that burn injury leads to epigenetic and transcriptional reprogramming of innate immune cells. Splenic F4/80 macrophages were isolated from mice at days 2, 9, and 14 days post-20% contact burn injury. Targeted transcriptomics and MAPit single-molecule chromatin profiling were used to assess immune, metabolic, and epigenetic changes. Canonical pathway analysis was performed to infer functional shifts over time. Burn injury induced a biphasic response in macrophages. Early after injury (Day 2), there was broad transcriptional suppression and epigenetic silencing of inflammatory regulators, including Stat3, Traf6, and Nfkb1. Over time (Days 9 and 14), loci associated with anti-inflammatory mediators such as Il-10 and Socs3 exhibited progressive chromatin opening and transcriptional upregulation. Metabolic gene profiles revealed persistent suppression of mitochondrial and oxidative phosphorylation programs. Canonical pathway analysis demonstrated early IL-10 signaling activation with sustained suppression of classical macrophage activation pathways. Chromatin architecture changes included nucleosome sliding and ejection events, consistent with dynamic, locus-specific regulation. This work challenges the classical notion of burn-induced immune suppression as purely a consequence of systemic inflammation. Instead, we reveal a programmed and locus-specific epigenetic architecture that may shape macrophage immune and metabolic function long after the acute phase.

immunology↗

bioRxiv2025

Clinical performance evaluation of a tiling amplicon panel for whole genome sequencing of respiratory syncytial virus

Accurate genomic characterization of respiratory syncytial virus (RSV) is crucial for studies of epidemiology and viral evolution, and monitoring potential escape from newly authorized vaccines and antivirals. We adapted a viral whole genome tiling amplicon panel (UW-ARTIC) and developed a custom bioinformatic pipeline for high-throughput, cost-effective sequencing of RSV-A and RSV-B. We established genome acceptability criteria and determined the performance characteristics of the panel including assay sensitivity, specificity, breadth of genome recovery, accuracy, and precision using contrived and remnant clinical specimens. High-quality genomes (>95% genome completeness; >500X and >1000X average depth for whole genome and fusion gene respectively) were recovered from samples with Ct [≤] 30 ([~]594 and 2,004 copies per reaction for RSV-A and RSV-B respectively). Minor variants were accurately identified in sample mixtures of 5:95 and higher. The assay showed high accuracy when compared against Sanger, shotgun metagenomic, and hybridization capture-based sequencing; and high repeatability and reproducibility. The UW-ARTIC RSV panel has utility in genomic surveillance, clinical and research applications. It has been used to generate FDA-reportable data for clinical trials of RSV antiviral products, with robust performance characteristics in samples from around the globe from as recently as the 2023/24 season. Continued genomic surveillance and future updates to primer sets will be essential for continued recovery of genomes as RSV continues to evolve.

genomics↗

bioRxiv2025

Minimization of gene editing off-target effects by tissue restriction of expression

Therapeutic in vivo gene editing with highly specific nucleases has the potential to revolutionize treatment for a wide range of human diseases, including genetic disorders and latent viral infections like herpes simplex virus (HSV). However, challenges regarding specificity, efficiency, delivery, and safety must be addressed before its clinical application. A key concern is the risk of off-target effects, which can cause unintended and potentially harmful genetic changes. We previously developed a curative in vivo gene editing approach to eliminate latent HSV using HSV-specific meganuclease delivered by an AAV vector. In this study, we investigate off-target effects of meganuclease by identifying potential off-target sites through GUIDE-tag analysis and assessing genetic alterations using amplicon deep sequencing in tissues from meganuclease treated mice. Our results show that meganuclease expression driven by a ubiquitous promoter leads to high off-target gene editing in the mouse liver, a non-relevant target tissue. However, restricting the meganuclease expression with a neuron-specific promoter and/or a liver-specific miRNA target sequence efficiently reduces off-target effects in both liver and trigeminal ganglia. These findings suggest that incorporation of regulatory DNA elements for tissue-specific expression in viral vectors can reduce off-target effects and improve the safety of therapeutic in vivo gene editing.

molecular biology↗

bioRxiv2020

Robust and sensitive detection of SARS-CoV-2 using PCR based methods

The World Health Organization (WHO) has declared the Coronavirus disease 2019 (COVID-19) as an international health emergency. Current diagnostic tests are based on the reverse transcription-quantitative polymerase chain reaction (RT-qPCR) method, the gold standard test that involves the amplification of viral RNA. However, the RT-qPCR assay has limitations in terms of sensitivity and quantification. In this study, we tested both qPCR and droplet digital PCR (ddPCR) to detect low amounts of viral RNA. The cycle threshold (CT) of viral RNA by RT-PCR significantly varied according to the sequence of primer and probe sets with in vitro transcript (IVT) RNA or viral RNA as templates, whereas the copy number of viral RNA by ddPCR was effectively quantified with IVT RNA, cultured viral RNA, and RNA from clinical samples. Furthermore, the clinical samples were assayed via both methods, and the sensitivity of the ddPCR was determined to be significantly higher than RT-qPCR. These findings suggest that ddPCR could be used as a highly sensitive and compatible diagnostic method for viral RNA detection.

microbiology↗

bioRxiv2020

Spike protein binding prediction with neutralizing antibodies of SARS-CoV-2

Coronavirus disease 2019 (COVID-19) is a new emerging human infectious disease caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2, also previously known as 2019-nCoV), originated in Wuhan seafood and animal market, China. Since December 2019, more than 69,000 cases of COVID-19 have been confirmed in China and quickly spreads to other counties. Currently, researchers put their best efforts to identify effective drugs for COVID-19. The neutralizing antibody, which binds to viral capsid in a manner that inhibits cellular entry of virus and uncoating of the genome, is the specific defense against viral invaders. In this study, we investigate to identify neutralizing antibodies that can bind to SARS-CoV-2 Sipke (S) protein and interfere with the interaction between viral S protein and a host receptor by bioinformatic methods. The sequence analysis of S protein showed two major differences in the RBD region of the SARS-CoV-2 S protein compared to SARS-CoV and SARS-CoV related bat viruses (btSARS-CoV). The insertion regions were close to interacting residues with the human ACE2 receptor. Epitope analysis of neutralizing antibodies revealed that SARS-CoV neutralizing antibodies used conformational epitopes, whereas MERS-CoV neutralizing antibodies used a common linear epitope region, which contributes to form the {beta}-sheet structure in MERS-CoV S protein and deleted in SARS-CoV-2 S protein. To identify effective neutralizing antibodies for SARS-CoV-2, the binding affinities of neutralizing antibodies with SARS-CoV-2 S protein were predicted and compared by antibody-antigen docking simulation. The result showed that CR3022 neutralizing antibody from human may have higher binding affinity with SARS-CoV-2 S protein than SARS-CoV S protein. We also found that F26G19 and D12 mouse antibodies could bind to SARS-CoV S protein with high affinity. Our findings provide crucial clues towards the development of antigen diagnosis, therapeutic antibody, and the vaccine against SARS-CoV-2.

bioinformatics↗

bioRxiv2020

Comparative analysis of primer-probe sets for the laboratory confirmation of SARS-CoV-2

Coronavirus disease 2019 (COVID-19) is newly emerging human infectious diseases, which is caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2, also previously known as 2019-nCoV). Within two months of the outbreak, more than 80,000 cases of COVID-19 have been confirmed worldwide. Since the human to human transmission occurred easily and the human infection is rapidly increasing, the sensitive and early diagnosis is essential to prevent the global outbreak. Recently, World Health Organization (WHO) announced various primer and probe sets for SARS-CoV-2 previously developed in China, Germany, Hong Kong, Japan, Thailand, and USA. In this study, we compared the ability to detect SARS-CoV-2 RNA among the seven primer-probe sets for N gene and the three primer-probe sets for Orf1 gene. The result of the comparative analysis represented that the 2019-nCoV_N2, N3 of USA and the ORF1ab of China are the most sensitive primer-probe sets for N and Orf1 genes, respectively. Therefore, the appropriate combination from ORF1ab (China), 2019-nCoV_N2, N3 (USA), and NIID_2019-nCOV_N (Japan) sets should be selected for the sensitive and reliable laboratory confirmation of SARS-CoV-2.

microbiology↗

bioRxiv2019

Metastatic function of METTL18 in breast cancer via actin methylation and Src

Recently, a SET domain containing 3 (SETD3) was identified as an actin histidine methyltransferase, functioning to control replication and pathogenesis in multiple mouse models for enterovirus infection as well as the regulation of smooth muscle contractility linked to primary dystocia. Here, in this study, we report another type of actin histidine methyltransferase, METTL18, that regulates the metastatic potential of breast cancer in human. Among methyltransferases, METTL18 was highly amplified in human breast cancer. In particular, poor prognosis was associated with high expression of METTL18 in HER2-negative breast cancer patients. This gene product was also found to be a critical component of metastatic responses. Loss of METTL18 expression significantly reduced metastatic responses of breast tumor cells both in vitro and in vivo. Mechanistically, it was observed that METTL18 increased actin polymerization, upregulated complex formation with HSP90AA1 and Src, enhanced the activity of an intermediate form of Src with tyrosine phosphorylation at both Y416 and Y527, and induced cellular metastatic responses, including morphological change, migration, and invasion of MDA-MB-231 cells in vitro and in mice. Methylated actin at His73 served as a critical site for interaction with HSP90AA1 and Src to activate p85/PI3K and STAT3. Our findings suggest that METTL18 plays critical roles in metastatic responses of HER2-negative breast cancer cells via actin polymerization and the generation of an intermediate form of Src.

cancer biology↗