• Journal of Life Science
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• v.29 no.9
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• pp.1034-1046
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• 2019

The mitochondria is the major cellular organelle of energy metabolism for the supply of cellular energy; it also plays an important role in controlling calcium regulation, reactive oxygen species (ROS) production, and apoptosis. Mitochondrial dysfunction causes various diseases, such as neurodegenerative diseases, Lou Gehrig's disease, cardiovascular disease, mental disorders, diabetes, and cancer. Most of the diseases are age-related diseases. In this review, we focus on the roles of mitochondrial dysfunction in cancer. Mitochondrial dysfunction induces carcinogenesis and is found in many cancers. The factors that cause mitochondrial dysfunction differ depending on the types of carcinoma, and those factors could cause cancer malignancy, such as resistance to therapy and metastasis. Mitochondrial dysfunction is caused by a lack of mitochondria, an inability to provide key substances, or a dysfunction in the ATP synthesis machinery. The main factor associated with cancer malignancy is mtDNA depletion. Mitochondrial dysfunction would leads to malignancy through changes in molecular activity or expression, but it is not known in detail which changes lead to cancer malignancy. In order to explore the relationship between mitochondrial dysfunction and cancer malignancy in detail, mitochondria dysfunctional cell lines are constructed using chemical methods such as EtBr treatment or gene editing methods, including shRNA and CRISPR/Cas9. Those mitochondria dysfunctional cell lines are used in the study of various diseases caused by mitochondrial dysfunction, including cancer.

• Journal of Plant Biotechnology
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• v.48 no.4
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• pp.207-222
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• 2021

Coffee is the most frequently consumed functional beverage world wide. The average daily coffee consumption is increasing. This crop, which plays an important role in the global economy is under great threat from climate change. To with stand the current climate change, farmers have to learn crop cultivation techniques, strategies to protect crops from diseases, and understand which type of seed varieties to use to avoid crop loss. The present review briefly discusses the coffee cultivation techniques, impact of climate changes on coffee production, processing techniques of coffee, and the importance of coffee in our society, including its chemical composition and prevention against, major diseases. Furthermore, the importance and role of advanced nanotechnology along with molecular approaches for coffee crop improvement and facing challenges are explained.

• Proceedings of the Korean Society of Crop Science Conference
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• 2019.04a
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• pp.137-137
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• 2019

• Molecules and Cells
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• v.43 no.5
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• pp.459-468
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• 2020

Nuclear receptor subfamily group H member 4 (NR1H4), also known as farnesoid X receptor, has been implicated in several cellular processes in the liver and intestine. Preclinical and clinical studies have suggested a role of NR1H4 in colon cancer development; however, how NR1H4 regulates colon cancer cell growth and survival remains unclear. We generated NR1H4 knockout (KO) colon cancer cells using clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein-9 nuclease (CAS9) technology and explored the effects of NR1H4 KO in colon cancer cell proliferation, survival, and apoptosis. Interestingly, NR1H4 KO cells showed impaired cell proliferation, reduced colony formation, and increased apoptotic cell death compared to control colon cancer cells. We identified MYC as an important mediator of the signaling pathway alterations induced by NR1H4 KO. NR1H4 silencing in colon cancer cells resulted in reduced MYC protein levels, while NR1H4 activation using an NR1H4 ligand, chenodeoxycholic acid, resulted in time- and dose-dependent MYC induction. Moreover, NR1H4 KO enhanced the anti-cancer effects of doxorubicin and cisplatin, supporting the role of MYC in the enhanced apoptosis observed in NR1H4 KO cells. Taken together, our findings suggest that modulating NR1H4 activity in colon cancer cells might be a promising alternative approach to treat cancer using MYC-targeting agents.

• Asian-Australasian Journal of Animal Sciences
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• v.31 no.4
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• pp.489-498
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• 2018

Objective: Foot and mouth disease (FMD) and porcine reproductive and respiratory syndrome (PRRS) are major diseases that interrupt porcine production. Because they are viral diseases, vaccinations are of only limited effectiveness in preventing outbreaks. To establish an alternative multi-resistant strategy against FMD virus (FMDV) and PRRS virus (PRRSV), the present study introduced two genetic modification techniques to porcine cells. Methods: First, cluster of differentiation 163 (CD163), the PRRSV viral receptor, was edited with the clustered regularly interspaced short palindromic repeats-CRISPR-associated protein 9 technique. The CD163 gene sequences of edited cells and control cells differed. Second, short hairpin RNA (shRNAs) were integrated into the cells. The shRNAs, targeting the 3D gene of FMDV and the open reading frame 7 (ORF7) gene of PRRSV, were transferred into fibroblasts. We also developed an in vitro shRNA verification method with a target gene expression vector. Results: shRNA activity was confirmed in vitro with vectors that expressed the 3D and ORF7 genes in the cells. Cells containing shRNAs showed lower transcript levels than cells with only the expression vectors. The shRNAs were integrated into CD163-edited cells to combine the two techniques, and the viral genes were suppressed in these cells. Conclusion: We established a multi-resistant strategy against viral diseases and an in vitro shRNA verification method.

• BMB Reports
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• v.52 no.2
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• pp.133-138
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• 2019

Upon viral infection, the 2', 5'-oligoadenylate synthetase (OAS)-ribonuclease L (RNaseL) system works to cleave viral RNA, thereby blocking viral replication. However, it is unclear whether OAS proteins have a role in regulating gene expression. Here, we show that OAS1 and OAS3 act as negative regulators of the expression of chemokines and interferon-responsive genes in human macrophages. Clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein-9 nuclease (Cas9) technology was used to engineer human myeloid cell lines in which the OAS1 or OAS3 gene was deleted. Neither OAS1 nor OAS3 was exclusively responsible for the degradation of rRNA in macrophages stimulated with poly(I:C), a synthetic surrogate for viral double-stranded (ds)RNA. An mRNA sequencing analysis revealed that genes related to type I interferon signaling and chemokine activity were increased in $OAS1^{-/-}$ and $OAS3^{-/-}$ macrophages treated with intracellular poly(I:C). Indeed, retinoic-acid-inducible gene (RIG)-I- and interferon-induced helicase C domain-containing protein (IFIH1 or MDA5)-mediated induction of chemokines and interferon-stimulated genes was regulated by OAS3, but Toll-like receptor 3 (TLR3)- and TLR4-mediated induction of those genes was modulated by OAS1 in macrophages. However, stimulation of these cells with type I interferons had no effect on OAS1- or OAS3-mediated chemokine secretion. These data suggest that OAS1 and OAS3 negatively regulate the expression of chemokines and interferon-responsive genes in human macrophages.

• Biomolecules & Therapeutics
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• v.30 no.2
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• pp.203-211
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• 2022

Melanogenesis is the production of melanin from tyrosine by a series of enzyme-catalyzed reactions, in which tyrosinase and DOPA oxidase play key roles. The melanin content in the skin determines skin pigmentation. Abnormalities in skin pigmentation lead to various skin pigmentation disorders. Recent research has shown that the expression of EMP2 is much lower in melanoma than in normal melanocytes, but its role in melanogenesis has not yet been elucidated. Therefore, we investigated the role of EMP2 in the melanogenesis of MNT1 human melanoma cells. We examined TRP-1, TRP-2, and TYR expression levels during melanogenesis in MNT1 melanoma cells by gene silencing of EMP2. Western blot and RT-PCR results confirmed that the expression levels of TYR and TRP-2 were decreased when EMP2 expression was knocked down by EMP2 siRNA in MNT1 cells, and these changes were reversed when EMP2 was overexpressed. We verified the EMP2 gene was knocked out of the cell line (EMP2 CRISPR/Cas9) by using a CRISPR/Cas9 system and found that the expression levels of TRP-2 and TYR were significantly lower in the EMP2 CRISPR/Cas9 cell lines. Loss of EMP2 also reduced migration and invasion of MNT1 melanoma cells. In addition, the melanosome transfer from the melanocytes to keratinocytes in the EMP2 KO cells cocultured with keratinocytes was reduced compared to the cells in the control coculture group. In conclusion, these results suggest that EMP2 is involved in melanogenesis via the regulation of TRP-2 expression.

• Research in Plant Disease
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• v.25 no.2
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• pp.49-61
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• 2019

Plant viruses are among the important pathogens that cause severe crop losses. The most efficient method to control viral diseases is currently to use virus resistant crops. In order to develop the virus resistant crops, a detailed understanding of the molecular interactions between viral and host proteins is necessary. Recessive resistance to a pathogen can be conferred when plant genes essential in the life cycle of a pathogens are deficient, while dominant resistance is mediated by host resistance (R) genes specifically interacting with effector proteins of pathogens. Thus, recessive resistance usually works more stably and broadly than dominant resistance. While most of the recessive resistance genes have so far been identified by forward genetic approaches, recent advances in genome editing technologies including CRISPR/Cas9 have increased interest in using these technologies as reverse genetic tools to engineer plant genes to confer recessive resistance. This review summarizes currently identified recessive resistance genes and introduces reverse genetic approaches to identify host interacting partner proteins of viral proteins and to evaluate the identified genes as genetic resources of recessive resistance. We further discuss recent advances in various precise genome editing technologies and how to apply these technologies to engineer plant immunity.

• Biomolecules & Therapeutics
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• v.29 no.5
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• pp.551-561
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• 2021

Thyroid cancer is the most common endocrine malignancy. Patients with well-differentiated thyroid cancers, such as papillary and follicular cancers, have a favorable prognosis. However, poorly differentiated thyroid cancers, such as medullary, squamous and anaplastic advanced thyroid cancers, are very aggressive and insensitive to radioiodine treatment. Thus, novel therapies that attenuate metastasis are urgently needed. We found that both PDGFC and PDGFRA are predominantly expressed in thyroid cancers and that the survival rate is significantly lower in patients with high PDGFRA expression. This finding indicates the important role of PDGF/PDGFR signaling in thyroid cancer development. Next, we established a SW579 squamous thyroid cancer cell line with 95.6% PDGFRA gene insertion and deletions (indels) through CRISPR/Cas9. Protein and invasion analysis showed a dramatic loss in EMT marker expression and metastatic ability. Furthermore, xenograft tumors derived from PDGFRA geneedited SW579 cells exhibited a minor decrease in tumor growth. However, distant lung metastasis was completely abolished upon PDGFRA gene editing, implying that PDGFRA could be an effective target to inhibit distant metastasis in advanced thyroid cancers. To translate this finding to the clinic, we used the most relevant multikinase inhibitor, imatinib, to inhibit PDGFRA signaling. The results showed that imatinib significantly suppressed cell growth, induced cell cycle arrest and cell death in SW579 cells. Our developed noninvasive apoptosis detection sensor (NIADS) indicated that imatinib induced cell apoptosis through caspase-3 activation. In conclusion, we believe that developing a specific and selective targeted therapy for PDGFRA would effectively suppress PDGFRA-mediated cancer aggressiveness in advanced thyroid cancers.

• Asian-Australasian Journal of Animal Sciences
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• v.30 no.8
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• pp.1183-1189
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• 2017

Objective: Owing to the public availability of complete genome sequences, including avian species, massive bioinformatics analyses may be conducted for computational gene prediction and the identification of gene regulatory networks through various informatics tools. However, to evaluate the biofunctional activity of a predicted target gene, in vivo and in vitro functional genomic analyses should be a prerequisite. Methods: Due to a lack of quail genomic sequence information, we first identified the partial genomic structure and sequences of the quail SH3 domain containing ring finger 2 (SH3RF2) gene. Subsequently, SH3RF2 was knocked out using clustered regularly interspaced short palindromic repeat/Cas9 technology and single cell-derived SH3RF2 mutant sublines were established to study the biofunctional activity of SH3RF2 in quail myoblast (QM7) cells during muscle differentiation. Results: Through a T7 endonuclease I assay and genotyping analysis, we established an SH3RF2 knockout (KO) QM7#4 subline with 61 and 155 nucleotide deletion mutations in SH3RF2. After the induction of myotube differentiation, the expression profiles were analyzed and compared between regular QM7 and SH3RF2 KO QM7#4 cells by global RNA sequencing and bioinformatics analysis. Conclusion: We did not detect any statistically significant role of SH3RF2 during myotube differentiation in QM7 myoblast cells. However, additional experiments are necessary to examine the biofunctional activity of SH3RF2 in cell proliferation and muscle growth.