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  • MinireviewSeptember 30, 2021

    1 1308 518

    Visualizing Live Chromatin Dynamics through CRISPR-Based Imaging Techniques

    Narendra Chaudhary , Jae-Kyeong Im , Si-Hyeong Nho , and Hajin Kim

    Mol. Cells 2021; 44(9): 627-636 https://doi.org/10.14348/molcells.2021.2254
    Abstract

    Abstract : The three-dimensional organization of chromatin and its time-dependent changes greatly affect virtually every cellular function, especially DNA replication, genome maintenance, transcription regulation, and cell differentiation. Sequencing-based techniques such as ChIP-seq, ATAC-seq, and Hi-C provide abundant information on how genomic elements are coupled with regulatory proteins and functionally organized into hierarchical domains through their interactions. However, visualizing the time-dependent changes of such organization in individual cells remains challenging. Recent developments of CRISPR systems for site-specific fluorescent labeling of genomic loci have provided promising strategies for visualizing chromatin dynamics in live cells. However, there are several limiting factors, including background signals, off-target binding of CRISPR, and rapid photobleaching of the fluorophores, requiring a large number of target-bound CRISPR complexes to reliably distinguish the target-specific foci from the background. Various modifications have been engineered into the CRISPR system to enhance the signal-to-background ratio and signal longevity to detect target foci more reliably and efficiently, and to reduce the required target size. In this review, we comprehensively compare the performances of recently developed CRISPR designs for improved visualization of genomic loci in terms of the reliability of target detection, the ability to detect small repeat loci, and the allowed time of live tracking. Longer observation of genomic loci allows the detailed identification of the dynamic characteristics of chromatin. The diffusion properties of chromatin found in recent studies are reviewed, which provide suggestions for the underlying biological processes.

  • Research ArticleSeptember 30, 2021

    2 1103 465

    An Essential Role of the N-Terminal Region of ACSL1 in Linking Free Fatty Acids to Mitochondrial β-Oxidation in C2C12 Myotubes

    Jinyan Nan , Ji Seon Lee , Seung-Ah Lee , Dong-Sup Lee , Kyong Soo Park , and Sung Soo Chung

    Mol. Cells 2021; 44(9): 637-646 https://doi.org/10.14348/molcells.2021.0077
    Abstract

    Abstract : Free fatty acids are converted to acyl-CoA by long-chain acyl-CoA synthetases (ACSLs) before entering into metabolic pathways for lipid biosynthesis or degradation. ACSL family members have highly conserved amino acid sequences except for their N-terminal regions. Several reports have shown that ACSL1, among the ACSLs, is located in mitochondria and mainly leads fatty acids to the β-oxidation pathway in various cell types. In this study, we investigated how ACSL1 was localized in mitochondria and whether ACSL1 overexpression affected fatty acid oxidation (FAO) rates in C2C12 myotubes. We generated an ACSL1 mutant in which the N-terminal 100 amino acids were deleted and compared its localization and function with those of the ACSL1 wild type. We found that ACSL1 adjoined the outer membrane of mitochondria through interaction of its N-terminal region with carnitine palmitoyltransferase-1b (CPT1b) in C2C12 myotubes. In addition, overexpressed ACSL1, but not the ACSL1 mutant, increased FAO, and ameliorated palmitate-induced insulin resistance in C2C12 myotubes. These results suggested that targeting of ACSL1 to mitochondria is essential in increasing FAO in myotubes, which can reduce insulin resistance in obesity and related metabolic disorders.

  • Research ArticleSeptember 30, 2021

    3 821 443

    Tumour-Derived Reg3A Educates Dendritic Cells to Promote Pancreatic Cancer Progression

    Jie Guo , Mengfan Liao , Xianmin Hu , and Jun Wang

    Mol. Cells 2021; 44(9): 647-657 https://doi.org/10.14348/molcells.2021.0145
    Abstract

    Abstract : As a pancreatic inflammatory marker, regenerating islet-derived protein 3A (Reg3A) plays a key role in inflammation-associated pancreatic carcinogenesis by promoting cell proliferation, inhibiting apoptosis, and regulating cancer cell migration and invasion. This study aimed to reveal a novel immuno-regulatory mechanism by which Reg3A modulates tumour-promoting responses during pancreatic cancer (PC) progression. In an in vitro Transwell system that allowed the direct co-culture of human peripheral blood-derived dendritic cells (DCs) and Reg3A-overexpressing/ silenced human PC cells, PC cell-derived Reg3A was found to downregulate CD80, CD83 and CD86 expression on educated DCs, increase DC endocytic function, inhibit DC-induced T lymphocyte proliferation, reduce IL-12p70 production, and enhance IL-23 production by DCs. The positive effect of tumour-derived Reg3A-educated human DCs on PC progression was demonstrated in vivo by intraperitoneally transferring them into PC-implanted severe combined immunodeficiency (SCID) mice reconstituted with human T cells. A Reg3A-JAK2/STAT3 positive feedback loop was identified in DCs educated with Reg3A. In conclusion, as a tumour-derived factor, Reg3A acted to block the differentiation and maturation of the most important antigen-presenting cells, DCs, causing them to limit their potential anti-tumour responses, thus facilitating PC escape and progression.

  • Research ArticleSeptember 30, 2021

    2 829 390

    Identification and Functional Characterization of Two Noncoding RNAs Transcribed from Putative Active Enhancers in Hepatocellular Carcinoma

    Ye-Eun Lee , Jiyeon Lee , Yong Sun Lee , Jiyoung Joan Jang , Hyeonju Woo , Hae In Choi , Young Gyu Chai , Tae-Kyung Kim , TaeSoo Kim , Lark Kyun Kim , and Sun Shim Choi

    Mol. Cells 2021; 44(9): 658-669 https://doi.org/10.14348/molcells.2021.0173
    Abstract

    Abstract : Enhancers have been conventionally perceived as cis-acting elements that provide binding sites for trans-acting factors. However, recent studies have shown that enhancers are transcribed and that these transcripts, called enhancer RNAs (eRNAs), have a regulatory function. Here, we identified putative eRNAs by profiling and determining the overlap between noncoding RNA expression loci and eRNA-associated histone marks such as H3K27ac and H3K4me1 in hepatocellular carcinoma (HCC) cell lines. Of the 132 HCC-derived noncoding RNAs, 74 overlapped with the eRNA loci defined by the FANTOM consortium, and 65 were located in the proximal regions of genes differentially expressed between normal and tumor tissues in TCGA dataset. Interestingly, knockdown of two selected putative eRNAs, THUMPD3-AS1 and LINC01572, led to downregulation of their target mRNAs and to a reduction in the proliferation and migration of HCC cells. Additionally, the expression of these two noncoding RNAs and target mRNAs was elevated in tumor samples in the TCGA dataset, and high expression was associated with poor survival of patients. Collectively, our study suggests that noncoding RNAs such as THUMPD3-AS1 and LINC01572 (i.e., putative eRNAs) can promote the transcription of genes involved in cell proliferation and differentiation and that the dysregulation of these noncoding RNAs can cause cancers such as HCC.

  • Research ArticleSeptember 30, 2021

    5 673 365

    Synaptotagmin 5 Controls SYP132-VAMP721/722 Interaction for Arabidopsis Immunity to Pseudomonas syringae pv tomato DC3000

    Soohong Kim , Hyeran Kim , Keunchun Park , Da Jeong Cho , Mi Kyung Kim , Chian Kwon , and Hye Sup Yun

    Mol. Cells 2021; 44(9): 670-679 https://doi.org/10.14348/molcells.2021.0100
    Abstract

    Abstract : Vesicle-associated membrane proteins 721 and 722 (VAMP721/722) are secretory vesicle-localized arginine-conserved soluble N-ethylmaleimide-sensitive factor attachment protein receptors (R-SNAREs) to drive exocytosis in plants. They are involved in diverse physiological processes in plants by interacting with distinct plasma membrane (PM) syntaxins. Here, we show that synaptotagmin 5 (SYT5) is involved in plant defense against Pseudomonas syringae pv tomato (Pst) DC3000 by regulating SYP132-VAMP721/722 interactions. Calcium-dependent stimulation of in vitro SYP132-VAMP722 interaction by SYT5 and reduced in vivo SYP132-VAMP721/722 interaction in syt5 plants suggest that SYT5 regulates the interaction between SYP132 and VAMP721/722. We interestingly found that disease resistance to Pst DC3000 bacterium but not to Erysiphe pisi fungus is compromised in syt5 plants. Since SYP132 plays an immune function to bacteria, elevated growth of surface-inoculated Pst DC3000 in VAMP721/722-deficient plants suggests that SYT5 contributes to plant immunity to Pst DC3000 by promoting the SYP132-VAMP721/722 immune secretory pathway.

  • Research ArticleSeptember 30, 2021

    4 775 387

    Regional TMPRSS2 V197M Allele Frequencies Are Correlated with COVID-19 Case Fatality Rates

    Sungwon Jeon , Asta Blazyte , Changhan Yoon , Hyojung Ryu , Yeonsu Jeon , Youngjune Bhak , Dan Bolser , Andrea Manica , Eun-Seok Shin , Yun Sung Cho , Byung Chul Kim , Namhee Ryoo , Hansol Choi , and Jong Bhak

    Mol. Cells 2021; 44(9): 680-687 https://doi.org/10.14348/molcells.2021.2249
    Abstract

    Abstract : Coronavirus disease, COVID-19 (coronavirus disease 2019), caused by SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2), has a higher case fatality rate in European countries than in others, especially East Asian ones. One potential explanation for this regional difference is the diversity of the viral infection efficiency. Here, we analyzed the allele frequencies of a nonsynonymous variant rs12329760 (V197M) in the TMPRSS2 gene, a key enzyme essential for viral infection and found a significant association between the COVID-19 case fatality rate and the V197M allele frequencies, using over 200,000 present-day and ancient genomic samples. East Asian countries have higher V197M allele frequencies than other regions, including European countries which correlates to their lower case fatality rates. Structural and energy calculation analysis of the V197M amino acid change showed that it destabilizes the TMPRSS2 protein, possibly negatively affecting its ACE2 and viral spike protein processing.

  • Research ArticleSeptember 30, 2021

    1 706 392

    Asunaprevir, a Potent Hepatitis C Virus Protease Inhibitor, Blocks SARS-CoV-2 Propagation

    Yun-Sook Lim , Lap P. Nguyen , Gun-Hee Lee , Sung-Geun Lee , Kwang-Soo Lyoo , Bumseok Kim , and Soon B. Hwang

    Mol. Cells 2021; 44(9): 688-695 https://doi.org/10.14348/molcells.2021.0076
    Abstract

    Abstract : The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has become a global health concern. Various SARS-CoV-2 vaccines have been developed and are being used for vaccination worldwide. However, no therapeutic agents against coronavirus disease 2019 (COVID-19) have been developed so far; therefore, new therapeutic agents are urgently needed. In the present study, we evaluated several hepatitis C virus direct-acting antivirals as potential candidates for drug repurposing against COVID-19. Theses include asunaprevir (a protease inhibitor), daclatasvir (an NS5A inhibitor), and sofosbuvir (an RNA polymerase inhibitor). We found that asunaprevir, but not sofosbuvir and daclatasvir, markedly inhibited SARS-CoV-2-induced cytopathic effects in Vero E6 cells. Both RNA and protein levels of SARS-CoV-2 were significantly decreased by treatment with asunaprevir. Moreover, asunaprevir profoundly decreased virion release from SARS-CoV-2-infected cells. A pseudoparticle entry assay revealed that asunaprevir blocked SARS-CoV-2 infection at the binding step of the viral life cycle. Furthermore, asunaprevir inhibited SARS-CoV-2 propagation in human lung Calu-3 cells. Collectively, we found that asunaprevir displays broad-spectrum antiviral activity and therefore might be worth developing as a new drug repurposing candidate for COVID-19.

  • Journal ClubSeptember 30, 2021

    1 372 170

    Sure, Fathers Give Birth, Too!

    Postnatal paternal folate deficiency increases congenital disabilities through H3K4me3
    histone methylation changes in sperm and embryos.

    Sun-Kyung Lee *

    Mol. Cells 2021; 44(9): 696-698 https://doi.org/10.14348/molcells.2021.0202
Mol. Cells
Sep 30, 2022 Vol.45 No.9
COVER PICTURE
The Target of Rapamycin Complex (TORC) is a central regulatory hub in eukaryotes, which is well conserved in diverse plant species, including tomato (Solanum lycopersicum). Inhibition of TORC genes (SlTOR, SlLST8, and SlRAPTOR) by VIGS (virus-induced gene silencing) results in early fruit ripening in tomato. The red/ orange tomatoes are early-ripened TORC-silenced fruits, while the green tomato is a control fruit. Top, left, control fruit (TRV2-myc); top, right, TRV2-SlLST8; bottom, left, TRV2-SlTOR; bottom, right, TRV2-SlRAPTOR(Choi et al., pp. 660-672).

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