Journal of Microbiology

Research Article

Efficiency of reverse genetics methods for rescuing severe acute respiratory syndrome coronavirus 2: Chang-Joo Park, Taehun Kim, Seung-Min Yoo, Myung-Shin Lee, Nam-Hyuk Cho, Changhoon Park; J. Microbiol. 2025;63(2):e2411023. Published online February 27, 2025; DOI: https://doi.org/10.71150/jm.2411023

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Abstract PDF: Bacteria-free reverse genetics techniques are crucial for the efficient generation of recombinant viruses, bypassing the need for labor-intensive bacterial cloning. These methods are particularly relevant for studying the pathogenesis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19. This study compared the efficiency of three bacteria-free approaches—circular polymerase extension reaction (CPER) with and without nick sealing and infectious sub-genomic amplicons (ISA)—to bacterial artificial chromosome (BAC)-based technology for rescuing SARS-CoV-2. Significant differences in viral titers following transfection were observed between methods. CPER with nick sealing generated virus titers comparable to those of the BAC-based method and 10 times higher than those of the standard CPER. In contrast, ISA demonstrated extremely low efficiency, as cytopathic effects were detected only after two passages. All rescued viruses exhibited replication kinetics consistent with those of the original strain, with no significant deviation in replication capacity. Furthermore, the utility of CPER and ISA in genetically modifying SARS-CoV-2 was demonstrated by successfully inserting the gene encoding green fluorescent protein into the genome. Overall, this study underscores the potential of bacteria-free methods, such as CPER and ISA, in advancing SARS-CoV-2 research while highlighting their significant differences in efficiency.

Journal Articles

Infection Dynamics of Dengue Virus in Caco-2 Cells Depending on Its Differentiation Status: Jayoung Nam, Jisu Lee, Geon A Kim, Seung-Min Yoo, Changhoon Park, Myung-Shin Lee; J. Microbiol. 2024;62(9):799-809. Published online August 30, 2024; DOI: https://doi.org/10.1007/s12275-024-00161-w

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Abstract: Dengue virus (DENV), from the Flaviviridae family, is the causative agent of dengue fever and poses a significant global health challenge. The virus primarily affects the vascular system and liver; however, a growing body of evidence suggests its involvement in the gastrointestinal (GI) tract, contributing to clinical symptoms such as abdominal pain, vomiting, and diarrhea. However, the mechanisms underlying DENV infection in the digestive system remain largely unexplored. Prior research has detected viral RNA in the GI tissue of infected animals; however, whether the dengue virus can directly infect human enterocytes remains unclear. In this study, we examine the infectivity of human intestinal cell lines to the dengue virus and their subsequent response. We report that the Caco-2 cell line, a model of human enterocytes, is susceptible to infection and capable of producing viruses. Notably, differentiated Caco-2 cells exhibited a lower infection rate yet a higher level of virus production than their undifferentiated counterparts. These findings suggest that human intestinal cells are a viable target for the dengue virus, potentially elucidating the GI symptoms observed in dengue fever and offering a new perspective on the pathogenetic mechanisms of the virus.

Rab27b regulates extracellular vesicle production in cells infected with Kaposi’s sarcoma–associated herpesvirus to promote cell survival and persistent infection: Hyungtaek Jeon , Su-Kyung Kang , Myung-Ju Lee , Changhoon Park , Seung-Min Yoo , Yun Hee Kang , Myung-Shin Lee; J. Microbiol. 2021;59(5):522-529. Published online April 20, 2021; DOI: https://doi.org/10.1007/s12275-021-1108-6

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Abstract

Extracellular vesicles (EVs) play a crucial role in cell-to-cell communication. EVs and viruses share several properties related to their structure and the biogenesis machinery in cells. EVs from virus-infected cells play a key role in virus spread and suppression using various loading molecules, such as viral proteins, host proteins, and microRNAs. However, it remains unclear how and why viruses regulate EV production inside host cells. The purpose of this study is to investigate the molecular mechanisms underlying EV production and their roles in Kaposi’s sarcoma-associated herpesvirus (KSHV)-infected cells. Here, we found that KSHV induced EV production in human endothelial cells via Rab- 27b upregulation. The suppression of Rab27b expression in KSHV-infected cells enhanced cell death by increasing autophagic flux and autolysosome formation. Our results indicate that Rab27b regulates EV biogenesis to promote cell survival and persistent viral infection during KSHV infection, thereby providing novel insights into the crucial role of Rab- 27b in the KSHV life cycle.

Citations

Citations to this article as recorded by

Engineered small extracellular vesicles as a novel platform to suppress human oncovirus-associated cancers
Iman Owliaee, Mehran khaledian, Armin Khaghani Boroujeni, Ali Shojaeian
Infectious Agents and Cancer.2023;[Epub] CrossRef
HMGB1, a potential regulator of tumor microenvironment in KSHV-infected endothelial cells
Myung-Ju Lee, Joohee Park, Seokjoo Choi, Seung-Min Yoo, Changhoon Park, Hong Seok Kim, Myung-Shin Lee
Frontiers in Microbiology.2023;[Epub] CrossRef
Alpha-2-macroglobulin as a novel diagnostic biomarker for human bladder cancer in urinary extracellular vesicles
Jisu Lee, Hyun Sik Park, Seung Ro Han, Yun Hee Kang, Ji Young Mun, Dong Wook Shin, Hyun-Woo Oh, Yoon-Kyoung Cho, Myung-Shin Lee, Jinsung Park
Frontiers in Oncology.2022;[Epub] CrossRef
Long non-coding RNAs in Sus scrofa ileum under starvation stress
Shu Wang, Yi Jia Ma, Yong Shi Li, Xu Sheng Ge, Chang Lu, Chun Bo Cai, Yang Yang, Yan Zhao, Guo Ming Liang, Xiao Hong Guo, Guo Qing Cao, Bu Gao Li, Peng Fei Gao
Animal Bioscience.2022; 35(7): 975. CrossRef

Latent Kaposi’s sarcoma-associated herpesvirus infection in bladder cancer cells promotes drug resistance by reducing reactive oxygen species: Suhyuk Lee , Jaehyuk Jang , Hyungtaek Jeon , Jisu Lee , Seung-Min Yoo , Jinsung Park , Myung-Shin Lee; J. Microbiol. 2016;54(11):782-788. Published online October 29, 2016; DOI: https://doi.org/10.1007/s12275-016-6388-x

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Abstract

Kaposi’s sarcoma-associated herpesvirus (KSHV) is the major etiologic agent of Kaposi’s sarcoma, primary effusion lymphoma, and multicentric Castleman’s disease. Recent studies have indicated that KSHV can be detected at high frequency in patient-derived bladder cancer tissue and might be associated with the pathogenesis of bladder cancer. Bladder cancer is the second most common cancer of the genitourinary tract, and it has a high rate of recurrence. Because drug resistance is closely related to chemotherapy failure and cancer recurrence, we investigated whether KSHV infection is associated with drug resistance of bladder cancer cells. Some KSHV-infected bladder cancer cell lines showed resistance to an anti-cancer drug, cisplatin, possibly as a result of downregulation of reactive oxygen species. Additionally, drug resistance acquired from KSHV infection could partly be overcome by HDAC1 inhibitors. Taken together, the data suggest the possible role of KSHV in chemo-resistant bladder cancer, and indicate the therapeutic potential of HDAC1 inhibitors in drug-resistant bladder cancers associated with KSHV infection.

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mBio.2024;[Epub] CrossRef
Genomic analysis of schistosomiasis-associated colorectal cancer reveals a unique mutational landscape and therapeutic implications
Dong Yu, Anqi Wang, Jing Zhang, Xinxing Li, Caifeng Jiang, Haiyang Zhou
Genes & Diseases.2023; 10(3): 657. CrossRef
Revisiting Histone Deacetylases in Human Tumorigenesis: The Paradigm of Urothelial Bladder Cancer
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Primary lymphocyte infection models for KSHV and its putative tumorigenesis mechanisms in B cell lymphomas
Sangmin Kang, Jinjong Myoung
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Research Support, Non-U.S. Gov't

NOTE] Kaposi’s Sarcoma-Associated Herpesvirus Infection of Endothelial Progenitor Cells Impairs Angiogenic Activity In Vitro: Seungchul Yoo , Sil Kim , Seungmin Yoo , In-Taek Hwang , Haewol Cho , Myung-Shin Lee; J. Microbiol. 2011;49(2):299-304. Published online May 3, 2011; DOI: https://doi.org/10.1007/s12275-011-0408-7

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Abstract: A recent study reported that endothelial progenitor cells (EPCs) are one of the reservoirs of Kaposi’s sarcoma associated herpesvirus (KSHV). Although EPCs are closely linked to angiogenesis and vasculogenesis, little is known about the angiogenic potential of KSHV in EPCs. In this study, we used EPCs isolated from human umbilical cord blood to show that early infection by KSHV in vitro impaired the neovascularization of EPCs in matrigel. Our results suggest that KSHV may disrupt the angiogenic potential of EPCs and that the disseminated infection of KSHV could be associated with EPC dysfunction.

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