197. Canine Adenovirus Type I (CAV-1) Infection of Human Cell-Lines

AbstractThe PML (promyelocytic leukemia) protein is a member of the TRIM family, a large group of proteins that show high diversity in functions but possess a common tripartite motif giving the family its name. We and others recently reported that both murine PML (mPML) and human PML (hPML) strongly restrict the early stages of infection by HIV-1 and other lentiviruses when expressed in mouse embryonic fibroblasts (MEFs). This restriction activity was found to contribute to the type I interferon (IFN-I)-mediated inhibition of HIV-1 in MEFs. Additionally, PML caused transcriptional repression of the HIV-1 promoter in MEFs. By contrast, the modulation of the early stages of HIV-1 infection of human cells by PML has been investigated by RNAi with unclear results. In order to conclusively determine whether PML restricts HIV-1 or not in human cells, we used CRISPR-Cas9 to knock out its gene in epithelial, lymphoid and monocytic human cell lines. Infection challenges showed that PML knockout had no effect on the permissiveness of these cells to HIV-1 infection. IFN-I treatments inhibited HIV-1 equally whether PML was expressed or not. Over-expression of individual hPML isoforms, or of mPML, in a human T cell line did not restrict HIV-1. The presence of PML was not required for the restriction of nonhuman retroviruses by TRIM5α was inhibited by arsenic trioxide through a PML-independent mechanism. We conclude that PML is not a restriction factor for HIV-1 in human cell lines representing diverse lineages.ImportancePML is involved in innate immune mechanisms against both DNA and RNA viruses. Although the mechanism by which PML inhibits highly divergent viruses is unclear, it was recently found that it can increase the transcription of interferon-stimulated genes (ISGs). However, whether human PML inhibits HIV-1 has been debated. Here we provide unambiguous, knockout-based evidence that PML does not restrict the early post-entry stages of HIV-1 infection in a variety of human cell types and does not participate in the inhibition of HIV-1 by IFN-I. Although this study does not exclude the possibility of other mechanisms by which PML may interfere with HIV-1, we nonetheless demonstrate that PML does not generally act as an HIV-1 restriction factor in human cells and that its presence is not required for IFN-I to stimulate the expression of anti-HIV-1 genes. These results contribute to uncovering the landscape of HIV-1 inhibition by ISGs in human cells.

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Gene Knockout Shows That PML (TRIM19) Does Not Restrict the Early Stages of HIV-1 Infection in Human Cell Lines

mSphere ◽

10.1128/mspheredirect.00233-17 ◽

2017 ◽

Vol 2 (3) ◽

Cited By ~ 4

Author(s):

Nasser Masroori ◽

Pearl Cherry ◽

Natacha Merindol ◽

Jia-xin Li ◽

Caroline Dufour ◽

...

Keyword(s):

Cell Lines ◽

Human Cell ◽

Gene Knockout ◽

Human Cells ◽

Restriction Factor ◽

Type I ◽

Human Cell Lines ◽

Early Stages ◽

Human T Cell ◽

Hiv 1

ABSTRACTThe PML (promyelocytic leukemia) protein is a member of the TRIM family, a large group of proteins that show high diversity in functions but possess a common tripartite motif giving the family its name. We and others recently reported that both murine PML (mPML) and human PML (hPML) strongly restrict the early stages of infection by HIV-1 and other lentiviruses when expressed in mouse embryonic fibroblasts (MEFs). This restriction activity was found to contribute to the type I interferon (IFN-I)-mediated inhibition of HIV-1 in MEFs. Additionally, PML caused transcriptional repression of the HIV-1 promoter in MEFs. In contrast, the modulation of the early stages of HIV-1 infection of human cells by PML has been investigated by RNA interference, with unclear results. In order to conclusively determine whether PML restricts HIV-1 or not in human cells, we used the clustered regularly interspaced short palindromic repeat with Cas9 (CRISPR-Cas9) system to knock out its gene in epithelial, lymphoid, and monocytic human cell lines. Infection challenges showed that PML knockout had no effect on the permissiveness of these cells to HIV-1 infection. IFN-I treatments inhibited HIV-1 equally whether PML was expressed or not. Overexpression of individual hPML isoforms, or of mPML, in a human T cell line did not restrict HIV-1. The presence of PML was not required for the restriction of nonhuman retroviruses by TRIM5α (another human TRIM protein), and TRIM5α was inhibited by arsenic trioxide through a PML-independent mechanism. We conclude that PML is not a restriction factor for HIV-1 in human cell lines representing diverse lineages.IMPORTANCEPML is involved in innate immune mechanisms against both DNA and RNA viruses. Although the mechanism by which PML inhibits highly divergent viruses is unclear, it was recently found that it can increase the transcription of interferon-stimulated genes (ISGs). However, whether human PML inhibits HIV-1 has been debated. Here we provide unambiguous, knockout-based evidence that PML does not restrict the early postentry stages of HIV-1 infection in a variety of human cell types and does not participate in the inhibition of HIV-1 by IFN-I. Although this study does not exclude the possibility of other mechanisms by which PML may interfere with HIV-1, we nonetheless demonstrate that PML does not generally act as an HIV-1 restriction factor in human cells and that its presence is not required for IFN-I to stimulate the expression of anti-HIV-1 genes. These results contribute to uncovering the landscape of HIV-1 inhibition by ISGs in human cells.

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SARS-CoV-2 disrupts proximal elements in the JAK-STAT pathway

Journal of Virology ◽

10.1128/jvi.00862-21 ◽

2021 ◽

Author(s):

Da-Yuan Chen ◽

Nazimuddin Khan ◽

Brianna J. Close ◽

Raghuveera K. Goel ◽

Benjamin Blum ◽

...

Keyword(s):

Cell Lines ◽

Human Cell ◽

Type I Interferon ◽

Cell Types ◽

Janus Kinase ◽

Type I ◽

Human Cell Lines ◽

Interferon Signaling ◽

Interferon Receptor ◽

Stat Pathway

SARS-CoV-2 can infect multiple organs, including lung, intestine, kidney, heart, liver, and brain. The molecular details of how the virus navigates through diverse cellular environments and establishes replication are poorly defined. Here, we generated a panel of phenotypically diverse, SARS-CoV-2-infectable human cell lines representing different body organs and performed longitudinal survey of cellular proteins and pathways broadly affected by the virus. This revealed universal inhibition of interferon signaling across cell types following SARS-CoV-2 infection. We performed systematic analyses of the JAK-STAT pathway in a broad range of cellular systems, including immortalized cells and primary-like cardiomyocytes, and found that SARS-CoV-2 targeted the proximal pathway components, including Janus kinase 1 (JAK1), tyrosine kinase 2 (Tyk2), and the interferon receptor subunit 1 (IFNAR1), resulting in cellular desensitization to type I IFN. Detailed mechanistic investigation of IFNAR1 showed that the protein underwent ubiquitination upon SARS-CoV-2 infection. Furthermore, chemical Inhibition of JAK kinases enhanced infection of stem cell-derived cultures, indicating that the virus benefits from inhibiting the JAK-STAT pathway. These findings suggest that the suppression of interferon signaling is a mechanism widely used by the virus to evade antiviral innate immunity, and that targeting the viral mediators of immune evasion may help block virus replication in patients with COVID-19. IMPORTANCE SARS-CoV-2 can infect various organs in the human body, but the molecular interface between the virus and these organs remains unexplored. In this study, we generated a panel of highly infectable human cell lines originating from various body organs and employed these cells to identify cellular processes commonly or distinctly disrupted by SARS-CoV-2 in different cell types. One among the universally impaired processes was interferon signaling. Systematic analysis of this pathway in diverse culture systems showed that SARS-CoV-2 targets the proximal JAK-STAT pathway components, destabilizes the type I interferon receptor though ubiquitination, and consequently renders the infected cells resistant to type I interferon. These findings illuminate how SARS-CoV-2 can continue to propagate in different tissues even in the presence of a disseminated innate immune response.

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Inhibition of interleukin-1 type I receptor expression in human cell-lines by an antisense phosphorothioate oligodeoxynucleotide

International Journal of Immunopharmacology ◽

10.1016/0192-0561(96)84502-4 ◽

1996 ◽

Vol 18 (4) ◽

pp. 227-240 ◽

Cited By ~ 6

Author(s):

Loren Miraglia ◽

Thomas Geiger ◽

C.Frank Bennett ◽

Nicholas M. Dean

Keyword(s):

Cell Lines ◽

Human Cell ◽

Interleukin 1 ◽

Receptor Expression ◽

Type I ◽

Human Cell Lines

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Dengue virus regulates type I interferon signalling in a strain-dependent manner in human cell lines

Journal of General Virology ◽

10.1099/vir.0.2008/001594-0 ◽

2008 ◽

Vol 89 (12) ◽

pp. 3052-3062 ◽

Cited By ~ 37

Author(s):

Indira Umareddy ◽

Kin Fai Tang ◽

Subhash G. Vasudevan ◽

Shamala Devi ◽

Martin L. Hibberd ◽

...

Keyword(s):

Cell Lines ◽

Human Cell ◽

Immune Escape ◽

Alternative Mechanism ◽

Type I ◽

Dependent Manner ◽

Human Cell Lines ◽

Type I Ifn ◽

Dengue Disease ◽

Strain Dependent

Outbreaks of dengue disease are constant threats to tropical and subtropical populations but range widely in severity, from mild to haemorrhagic fevers, for reasons that are still elusive. We investigated the interferon (IFN) response in infected human cell lines A549 and HepG2, using two strains (NGC and TSV01) of dengue serotype 2 (DEN2) and found that the two viruses exhibited a marked difference in inducing type I IFN response. While TSV01 infection led to activation of type I antiviral genes such as EIF2AK2 (PKR), OAS, ADAR and MX, these responses were absent in NGC-infected cells. Biochemical analysis revealed that NGC but not TSV01 suppressed STAT-1 and STAT-2 activation in response to type I IFN (α and β). However, these two strains did not differ in their response to type II IFN (γ). Although unable to suppress IFN signalling, TSV01 infection caused a weaker IFN-β induction compared with NGC, suggesting an alternative mechanism of innate immune escape. We extended our study to clinical isolates of various serotypes and found that while MY10245 (DEN2) and MY22713 (DEN4) could suppress the IFN response in a similar fashion to NGC, three other strains of dengue [EDEN167 (DEN1), MY02569 (DEN1) and MY10340 (DEN2)] were unable to suppress the IFN response, suggesting that this difference is strain-dependent but not serotype-specific. Our report indicates the existence of a strain-specific virulence factor that may impact on disease severity.

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Usutu virus growth in human cell lines: induction of and sensitivity to type I and III interferons

Journal of General Virology ◽

10.1099/vir.0.046433-0 ◽

2013 ◽

Vol 94 (4) ◽

pp. 789-795 ◽

Cited By ~ 10

Author(s):

Carolina Scagnolari ◽

Beniamino Caputo ◽

Simona Trombetti ◽

Giulia Cacciotti ◽

Annalisa Soldà ◽

...

Keyword(s):

Cell Lines ◽

Human Cell ◽

Vero Cells ◽

Growth Cycle ◽

Type I ◽

Human Cell Lines ◽

Inhibitory Effects ◽

Virus Growth ◽

Usutu Virus ◽

Pre Treatment

The mechanisms of Usutu virus (USUV) pathogenesis are largely unknown. The aim of this study was to evaluate the sensitivity of USUV to interferon (IFN) and the capacity of USUV to stimulate IFN production. Initial experiments were conducted to characterize the susceptibility of human cell lines to USUV infection and to evaluate the single-growth cycle replication curve of USUV. Results indicate that USUV is able to infect a variety of human cell lines, completing the replication cycle in Hep-2 and Vero cells within 48 h. Pre-treatment of cells with types I and III IFNs significantly inhibited the replication of USUV. However, the inhibitory effects of IFNs were considerably less if IFN was added after viral infection had been initiated. Also, USUV weakly induced types I and III IFNs.

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