scholarly journals TRIM26 is a critical host factor for HCV replication and contributes to host tropism

2021 ◽  
Vol 7 (2) ◽  
pp. eabd9732
Author(s):  
Yisha Liang ◽  
Guigen Zhang ◽  
Qiheng Li ◽  
Lin Han ◽  
Xiaoyou Hu ◽  
...  
Keyword(s):  
Hepatoma Cell ◽  
Ectopic Expression ◽  
Host Factor ◽  
Hepatoma Cell Line ◽  
Genome Replication ◽  
Host Tropism ◽  
Host Interactions ◽  
Genome Wide ◽  
Factor Deficiency ◽  
A Genome

Hepatitis C virus (HCV) remains a major human pathogen that requires better understanding of virus-host interactions. In this study, we performed a genome-wide CRISPR-Cas9 screening and identified TRIM26, an E3 ligase, as a critical HCV host factor. Deficiency of TRIM26 specifically impairs HCV genome replication. Mechanistic studies showed that TRIM26 interacts with HCV-encoded NS5B protein and mediates its K27-linked ubiquitination at residue K51, and thus promotes the NS5B-NS5A interaction. Moreover, mouse TRIM26 does not support HCV replication because of its unique six–amino acid insert that prevents its interaction with NS5B. Ectopic expression of human TRIM26 in a mouse hepatoma cell line that has been reconstituted with other essential HCV host factors promotes HCV infection. In conclusion, we identified TRIM26 as a host factor for HCV replication and a new determinant of host tropism. These results shed light on HCV-host interactions and may facilitate the development of an HCV animal model.

scholarly journals Metabolic precision labeling enables selective probing of O-linkedN-acetylgalactosamine glycosylation

2020 ◽  
Vol 117 (41) ◽  
pp. 25293-25301
Author(s):  
Marjoke F. Debets ◽  
Omur Y. Tastan ◽  
Simon P. Wisnovsky ◽  
Stacy A. Malaker ◽  
Nikolaos Angelis ◽  
...  
Keyword(s):  
Cell Surface ◽  
Secretory Pathway ◽  
Ectopic Expression ◽  
Protein Glycosylation ◽  
Major Type ◽  
Metabolic Labeling ◽  
Uridine Diphosphate ◽  
Superresolution Microscopy ◽  
Genome Wide ◽  
A Genome

Protein glycosylation events that happen early in the secretory pathway are often dysregulated during tumorigenesis. These events can be probed, in principle, by monosaccharides with bioorthogonal tags that would ideally be specific for distinct glycan subtypes. However, metabolic interconversion into other monosaccharides drastically reduces such specificity in the living cell. Here, we use a structure-based design process to develop the monosaccharide probeN-(S)-azidopropionylgalactosamine (GalNAzMe) that is specific for cancer-relevant Ser/Thr(O)–linkedN-acetylgalactosamine (GalNAc) glycosylation. By virtue of a branchedN-acylamide side chain, GalNAzMe is not interconverted by epimerization to the correspondingN-acetylglucosamine analog by the epimeraseN-acetylgalactosamine–4-epimerase (GALE) like conventional GalNAc–based probes. GalNAzMe enters O-GalNAc glycosylation but does not enter other major cell surface glycan types including Asn(N)-linked glycans. We transfect cells with the engineered pyrophosphorylase mut-AGX1 to biosynthesize the nucleotide-sugar donor uridine diphosphate (UDP)-GalNAzMe from a sugar-1-phosphate precursor. Tagged with a bioorthogonal azide group, GalNAzMe serves as an O-glycan–specific reporter in superresolution microscopy, chemical glycoproteomics, a genome-wide CRISPR-knockout (CRISPR-KO) screen, and imaging of intestinal organoids. Additional ectopic expression of an engineered glycosyltransferase, “bump-and-hole” (BH)–GalNAc-T2, boosts labeling in a programmable fashion by increasing incorporation of GalNAzMe into the cell surface glycoproteome. Alleviating the need for GALE-KO cells in metabolic labeling experiments, GalNAzMe is a precision tool that allows a detailed view into the biology of a major type of cancer-relevant protein glycosylation.

scholarly journals Systematic Analysis of Gibberellin Pathway Components in Medicago truncatula Reveals the Potential Application of Gibberellin in Biomass Improvement

2020 ◽  
Vol 21 (19) ◽  
pp. 7180
Author(s):  
Hongfeng Wang ◽  
Hongjiao Jiang ◽  
Yiteng Xu ◽  
Yan Wang ◽  
Lin Zhu ◽  
...  
Keyword(s):  
Medicago Truncatula ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Ectopic Expression ◽  
Feedback Regulation ◽  
Specific Expression ◽  
Systematic Analysis ◽  
Model Legume ◽  
Genome Wide ◽  
A Genome

Gibberellins (GAs), a class of phytohormones, act as an essential natural regulator of plant growth and development. Many studies have shown that GA is related to rhizobial infection and nodule organogenesis in legume species. However, thus far, GA metabolism and signaling components are largely unknown in the model legume Medicago truncatula. In this study, a genome-wide analysis of GA metabolism and signaling genes was carried out. In total 29 components, including 8 MtGA20ox genes, 2 MtGA3ox genes, 13 MtGA2ox genes, 3 MtGID1 genes, and 3 MtDELLA genes were identified in M. truncatula genome. Expression profiles revealed that most members of MtGAox, MtGID1, and MtDELLA showed tissue-specific expression patterns. In addition, the GA biosynthesis and deactivation genes displayed a feedback regulation on GA treatment, respectively. Yeast two-hybrid assays showed that all the three MtGID1s interacted with MtDELLA1 and MtDELLA2, suggesting that the MtGID1s are functional GA receptors. More importantly, M. truncatula exhibited increased plant height and biomass by ectopic expression of the MtGA20ox1, suggesting that enhanced GA response has the potential for forage improvement.

scholarly journals AP4 is a mediator of epithelial–mesenchymal transition and metastasis in colorectal cancer

2013 ◽  
Vol 210 (7) ◽  
pp. 1331-1350 ◽  
Author(s):  
Rene Jackstadt ◽  
Simone Röh ◽  
Jens Neumann ◽  
Peter Jung ◽  
Reinhard Hoffmann ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Target Genes ◽  
Epithelial Mesenchymal Transition ◽  
Ectopic Expression ◽  
Migration And Invasion ◽  
Mesenchymal Transition ◽  
Helix Loop Helix ◽  
Genome Wide ◽  
A Genome ◽  
Tumor Types

The basic helix-loop-helix transcription factor AP4/TFAP4/AP-4 is encoded by a c-MYC target gene and displays up-regulation concomitantly with c-MYC in colorectal cancer (CRC) and numerous other tumor types. Here a genome-wide characterization of AP4 DNA binding and mRNA expression was performed using a combination of microarray, genome-wide chromatin immunoprecipitation, next-generation sequencing, and bioinformatic analyses. Thereby, hundreds of induced and repressed AP4 target genes were identified. Besides many genes involved in the control of proliferation, the AP4 target genes included markers of stemness (LGR5 and CD44) and epithelial–mesenchymal transition (EMT) such as SNAIL, E-cadherin/CDH1, OCLN, VIM, FN1, and the Claudins 1, 4, and 7. Accordingly, activation of AP4 induced EMT and enhanced migration and invasion of CRC cells. Conversely, down-regulation of AP4 resulted in mesenchymal–epithelial transition and inhibited migration and invasion. In addition, AP4 induction was required for EMT, migration, and invasion caused by ectopic expression of c-MYC. Inhibition of AP4 in CRC cells resulted in decreased lung metastasis in mice. Elevated AP4 expression in primary CRC significantly correlated with liver metastasis and poor patient survival. These findings imply AP4 as a new regulator of EMT that contributes to metastatic processes in CRC and presumably other carcinomas.

scholarly journals A Genome-wide CRISPR Screen Identifies ZCCHC14 as a Host Factor Required for Hepatitis B Surface Antigen Production

Cell Reports ◽  
2019 ◽  
Vol 29 (10) ◽  
pp. 2970-2978.e6 ◽  
Author(s):  
Anastasia Hyrina ◽  
Christopher Jones ◽  
Darlene Chen ◽  
Scott Clarkson ◽  
Nadire Cochran ◽  
...  
Keyword(s):  
Hepatitis B ◽  
Surface Antigen ◽  
Hepatitis B Surface Antigen ◽  
Host Factor ◽  
Antigen Production ◽  
Genome Wide ◽  
A Genome ◽  
Crispr Screen

scholarly journals Dual RNAseq shows the human mucosal immunity protein, MUC13, is a hallmark of Plasmodium exoerythrocytic infection

2017 ◽  
Author(s):  
Gregory M. LaMonte ◽  
Pamela Orjuela-Sanchez ◽  
Lawrence Wang ◽  
Shangzhong Li ◽  
Justine Swann ◽  
...  
Keyword(s):  
Mucosal Immunity ◽  
Hepatoma Cell ◽  
Critical Window ◽  
Hepatoma Cell Lines ◽  
Genome Wide ◽  
A Genome ◽  
Infected Cells ◽  
Parasite Replication ◽  
Host Parasite ◽  
Stage Infection

SummaryThe exoerythrocytic stage of Plasmodium malaria infection is a critical window for prophylactic intervention. Using a genome-wide dual RNA sequencing of flow-sorted infected and uninfected hepatoma cells we identify the human mucosal immunity gene, Mucin13 (MUC13), as strongly upregulated during Plasmodium exoerythrocytic hepatic-stage infection. We confirm that MUC13 expression is upregulated in hepatoma cell lines and primary hepatocytes. In immunofluorescence assays, host MUC13 protein expression distinguishes infected cells from adjacent uninfected cells and shows similar colocalization with parasite biomarkers such as UIS4 and HSP70. We further show that localization patterns are species independent, distinguishing both P. berghei and P. vivax infected cells, and that MUC13 can be used to identify compounds that inhibit parasite replication in hepatocytes across all Human-infecting Plasmodium species. This data presents a novel interface of host-parasite interactions in Plasmodium, in that a component of host mucosal immunity is reprogrammed to assist the progression of infection.

scholarly journals TMEM41B is a host factor required for the replication of diverse coronaviruses including SARS-CoV-2

2021 ◽  
Vol 17 (5) ◽  
pp. e1009599
Author(s):  
Joseph D. Trimarco ◽  
Brook E. Heaton ◽  
Ryan R. Chaparian ◽  
Kaitlyn N. Burke ◽  
Raquel A. Binder ◽  
...  
Keyword(s):  
Host Factor ◽  
Host Protein ◽  
Loss Of Function ◽  
Host Proteins ◽  
Replication Complex ◽  
Genome Wide ◽  
Host Membrane ◽  
A Genome ◽  
Viral Replication Complex ◽  
Viral Lifecycle

Antiviral therapeutics are a front-line defense against virally induced diseases. Because viruses frequently mutate to escape direct inhibition of viral proteins, there is interest in targeting the host proteins that the virus must co-opt to complete its replication cycle. However, a detailed understanding of the interactions between the virus and the host cell is necessary in order to facilitate development of host-directed therapeutics. As a first step, we performed a genome-wide loss of function screen using the alphacoronavirus HCoV-229E to better define the interactions between coronaviruses and host-factors. We report the identification and validation of an ER-resident host protein, TMEM41B, as an essential host factor for not only HCoV-229E but also genetically distinct coronaviruses including the pandemic beta-coronavirus SARS-CoV-2. We show that the protein is required at an early, but post-receptor engagement, stage of the viral lifecycle. Further, mechanistic studies revealed that although the protein was not enriched at replication complexes, it likely contributes to viral replication complex formation via mobilization of cholesterol and other lipids to facilitate host membrane expansion and curvature. Continued study of TMEM41B and the development of approaches to prevent its function may lead to broad spectrum anti-coronavirus therapeutics.

scholarly journals Recombination events are concentrated in the spike protein region of Betacoronaviruses

PLoS Genetics ◽  
2020 ◽  
Vol 16 (12) ◽  
pp. e1009272
Author(s):  
Louis-Marie Bobay ◽  
Angela C. O’Donnell ◽  
Howard Ochman
Keyword(s):  
Experimental Studies ◽  
Host Cells ◽  
Spike Protein ◽  
Mutation Rates ◽  
Gene Exchange ◽  
Host Tropism ◽  
Genome Wide ◽  
A Genome ◽  
Genomic Regions ◽  
Sequence Similarities

The Betacoronaviruses comprise multiple subgenera whose members have been implicated in human disease. As with SARS, MERS and now SARS-CoV-2, the origin and emergence of new variants are often attributed to events of recombination that alter host tropism or disease severity. In most cases, recombination has been detected by searches for excessively similar genomic regions in divergent strains; however, such analyses are complicated by the high mutation rates of RNA viruses, which can produce sequence similarities in distant strains by convergent mutations. By applying a genome-wide approach that examines the source of individual polymorphisms and that can be tested against null models in which recombination is absent and homoplasies can arise only by convergent mutations, we examine the extent and limits of recombination in Betacoronaviruses. We find that recombination accounts for nearly 40% of the polymorphisms circulating in populations and that gene exchange occurs almost exclusively among strains belonging to the same subgenus. Although experimental studies have shown that recombinational exchanges occur at random along the coronaviral genome, in nature, they are vastly overrepresented in regions controlling viral interaction with host cells.

scholarly journals From bedside to bench: regulation of host factors in SARS-CoV-2 infection

2021 ◽  
Author(s):  
Samantha Y. Q. Ong ◽  
Ilya M. Flyamer ◽  
Wendy A. Bickmore ◽  
Simon C. Biddie
Keyword(s):  
Association Studies ◽  
Host Factor ◽  
Host Factors ◽  
Genome Wide Association Studies ◽  
Angiotensin Converting Enzyme 2 ◽  
Host Interactions ◽  
Genome Wide ◽  
Therapeutic Development ◽  
Transmembrane Serine Protease ◽  
Chromatin Biology

AbstractThe zoonotic coronavirus SARS-CoV-2 (severe acute respiratory syndrome coronavirus-2), which causes COVID-19 (coronavirus disease-2019), has resulted in a pandemic. This has led to an urgent need to understand the molecular determinants of SARS-CoV-2 infection, factors associated with COVID-19 heterogeneity and severity, and therapeutic options for these patients. In this review, we discuss the role of host factors in SARS-CoV-2 infection and describe variations in host factor expression as mechanisms underlying the symptoms and severity of COVID-19. We focus on two host factors, angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2), implicated in SARS-CoV-2 infection. We also discuss genetic variants associated with COVID-19 severity revealed in selected patients and based on genome-wide association studies (GWASs). Furthermore, we highlight important advances in cell and chromatin biology, such as single-cell RNA and chromatin sequencing and chromosomal conformation assays, as methods that may aid in the discovery of viral–host interactions in COVID-19. Understanding how regulation of host factor genes varies in physiological and pathological states might explain the heterogeneity observed in SARS-CoV-2 infection, help identify pathways for therapeutic development, and identify patients most likely to progress to severe COVID-19.

scholarly journals SATB2 Overexpression Promotes the Proliferation, Migration and Invasion of Oral Squamous Cell Carcinoma by Up-Regulating NOX4

2020 ◽  
Author(s):  
Jie Gao ◽  
Ying Meng ◽  
Xin Ge ◽  
Chen-xing Hou ◽  
Qing-hai Zhu ◽  
...  
Keyword(s):  
Squamous Cell Carcinoma ◽  
Oral Squamous Cell Carcinoma ◽  
Cell Carcinoma ◽  
Squamous Cell ◽  
Ectopic Expression ◽  
Migration And Invasion ◽  
Clinical Samples ◽  
Genome Wide ◽  
A Genome ◽  
Sirna Knockdown

Abstract Background: While atypical expression of special AT-rich sequence-binding protein 2 (SATB2) has been approved associated with tumor progression, metastasis and unfavourable prognosis in various carcinomas. However, in oral squamous cell carcinoma (OSCC), both the expressive state and associated functions of SATB2’s are still undefined.Methods: Real-time PCR, western blotting, and immunohistochemistry were used to examine SATB2 expression. In vitro experiments including Flow Cytometry, CCK8 assay, migration assay, wound-healing assay were used to investigate the effects of SATB2 on HN4 cell proliferation, migration and invasion ability. Additionally, an orthotopic implantation assay was performed in nude mice to confirm the effects of SATB2 in vivo. Furthermore, a genome wide siRNA knockdown experiment was performed to explore the potential downstream regulatory mechanism of SATB2 in OSCC.Results: We found that , in clinical samples from a retrospective cohort of 58 OSCC patients, high expression of SATB2 is associated with poor prognosis of OSCC patients. In this study, we investigated SATB2 is highly expressed in OSCC tissues and cell lines ,which can promotes OSCC cells’ proliferation, migration, invasion and tumor growth. Following a genome wide siRNA knockdown experiment, we identified NOX4, a bona fide downstream target of SATB2, which can partially suppress OSCC proliferation. Furthermore, NOX4 knockdown inhibits tumorigenicity, which can be rescued partially by ectopic expression of SATB2.Conclusion: Our findings not only indicate overexpression of SATB2 triggers the proliferative, migratory and invasive mechanisms which are important in the malignant phenotype of OSCC, but also identify NOX4 as the downstream gene for SATB2. These findings indicate that SATB2 may play a key role in OSCC tumorigenicity and may be a future target for the development of new therapeutic regimens.

scholarly journals Genome-wide CRISPR screen identifies RACK1 as a critical host factor for flavivirus replication

2021 ◽  
Author(s):  
Byron Shue ◽  
Abhilash I. Chiramel ◽  
Berati Cerikan ◽  
Thu-Hien To ◽  
Sonja Frölich ◽  
...  
Keyword(s):  
Viral Replication ◽  
Host Factor ◽  
Host Protein ◽  
Ns1 Protein ◽  
Genome Wide ◽  
A Genome ◽  
Wide Range ◽  
Cellular Factors ◽  
Novel Host ◽  
The Impact

Cellular factors have important roles in all facets of the flavivirus replication cycle. Deciphering viral-host protein interactions is essential for understanding the flavivirus lifecycle as well as development of effective antiviral strategies. To uncover novel host factors that are co-opted by multiple flaviviruses, a CRISPR/Cas9 genome wide knockout (KO) screen was employed to identify genes required for replication of Zika virus (ZIKV). Receptor for Activated Protein C Kinase 1 (RACK1) was identified as a novel host factor required for ZIKV replication, which was confirmed via complementary experiments. Depletion of RACK1 via siRNA demonstrated that RACK1 is important for replication of a wide range of mosquito- and tick-borne flaviviruses, including West Nile Virus (WNV), Dengue Virus (DENV), Powassan Virus (POWV) and Langat Virus (LGTV) as well as the coronavirus SARS-CoV-2, but not for YFV, EBOV, VSV or HSV. Notably, flavivirus replication was only abrogated when RACK1 expression was dampened prior to infection. Utilising a non-replicative flavivirus model, we show altered morphology of viral replication factories and reduced formation of vesicle packets (VPs) in cells lacking RACK1 expression. In addition, RACK1 interacted with NS1 protein from multiple flaviviruses; a key protein for replication complex formation. Overall, these findings reveal RACK1’s crucial role to the biogenesis of pan-flavivirus replication organelles. Importance Cellular factors are critical in all facets of viral lifecycles, where overlapping interactions between the virus and host can be exploited as possible avenues for the development of antiviral therapeutics. Using a genome-wide CRISPR knock-out screening approach to identify novel cellular factors important for flavivirus replication we identified RACK1 as a pro-viral host factor for both mosquito- and tick-borne flaviviruses in addition to SARS-CoV-2. Using an innovative flavivirus protein expression system, we demonstrate for the first time the impact of the loss of RACK1 on the formation of viral replication factories known as 'vesicle packets' (VPs). In addition, we show that RACK1 can interact with numerous flavivirus NS1 proteins as a potential mechanism by which VP formation can be induced by the former.

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