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Open Access Publications from the University of California

Scholarly Works (6 results)

  • Thesis
  • Peer Reviewed
UCLA Electronic Theses and Dissertations (2012)

Interferons (IFNs) are essential anti-viral cytokines that suppress viral infections by upregulation of antiviral effectors, collectively called interferon-stimulated genes (ISGs). We identified ISGs by gene expression profiling and performed functional screen of 288 ISGs that have antiviral activity against vesicular stomatitis virus (VSV) and murine gammaherpes virus (MHV68). Validation of antiviral genes from the screen demonstrated ISGs have inhibitory effect on specific virus. In particular, BMP2 inhibited MHV68 but not VSV replication. Conversely, Tap1 overexpression and deficiency led to resistance and susceptibility to VSV infection, respectively, while having effect on MHV68 growth.

Initially identified by its antiviral activity against VSV and MHV68, cholesterol-25-hydroxylase (Ch25h) was found to be an ISG that cause antiviral effect by production of a soluble oxysterol, 25-hydroxycholesterol. Toll-like receptor agonists induced Ch25h in an IFN-dependent manner. Ch25h overexpression and 25HC treatment broadly inhibited replication of other enveloped RNA and DNA viruses, including HIV, HSV, RSSEV, RVFV, and EBOV. Ch25h and 25HC inhibited efficiency of viral fusion in VSV and HIV. Administration of 25HC suppresses HIV replication in humanized mice, while functional loss of Ch25h lead to increased susceptibility to infections in vitro and in vivo.

Since interferons regulate many genes that can have profound impact on cellular homeostasis, regulation of IFN is tightly regulated. We have identified retinoid-X-receptors (RXR) to be a suppressor of type-I IFN response. Overexpression of RXR and activation of RXR with RXR-specific agonists led to increased viral susceptibility to viral infection in RAW264.7 cell lines. Functional loss of RXR in F9 embryonal caricoma cell lines led to increased resistance to viral infection. RXR agonists inhibited phosphorylation of upstream signaling activators of IFN, TBK and IRF3. Gene expression analyses show RXR inhibited many ISGs induced by dsRNA mimetic, polyI:C and partial suppression of genes induced by lipidA.

In effort to use virus as a tool to improve immunity, we developed an approach for live-attenuated vaccine development by genome-wide profiling. Using flu as a model, we generated high-density mutations in the influenza (WSN strain) M-gene and profiled the fitness of every mutation in vivo through time. By selecting mutants that exhibited an attenuated growth profile, we identified a mutant, W7-791, that grew in vitro, infected mice, and was not virulent. Vaccination of mice with W7-791 conferred protection against lethal infectious dose of two strains of H1N1 influenza and a phylogenetically disparate, H3N2 virus.

    Cover page: Effector Functions and Regulation of Host Antiviral Immunity
    • Article
    • Peer Reviewed
    UCLA Previously Published Works (2015)
    Rapid and robust induction of type I IFN (IFN-I) is a critical event in host antiviral innate immune response. It has been well demonstrated that cyclic GMP-AMP (cGAMP) synthase (cGAS) plays an important role in sensing cytosolic DNA and triggering STING dependent signaling to induce IFN-I. However, it is largely unknown how cGAS itself is regulated during pathogen infection and IFN-I production. In this study, we show that pattern recognition receptor (PRR) ligands, including lipid A, LPS, poly(I:C), poly(dA:dT), and cGAMP, induce cGAS expression in an IFN-I-dependent manner in both mouse and human macrophages. Further experiments indicated that cGAS is an IFN-stimulated gene (ISG), and two adjacent IFN-sensitive response elements (ISREs) in the promoter region of cGAS mediate the induction of cGAS by IFN-I. Additionally, we show that optimal production of IFN-β triggered by poly (dA:dT) or HSV-1 requires IFNAR signaling. Knockdown of the constitutively expressed DNA sensor DDX41 attenuates poly(dA:dT)-triggered IFN-β production and cGAS induction. By analyzing the dynamic expression of poly(dA:dT)-induced IFN-β and cGAS transcripts, we have found that induction of IFN-β is earlier than cGAS. Furthermore, we have provided evidence that induction of cGAS by IFN-I meditates the subsequent positive feedback regulation of DNA-triggered IFN-I production. Thus, our study not only provides a novel mechanism of modulating cGAS expression, but also adds another layer of regulation in DNA-triggered IFN-I production by induction of cGAS.
      Cover page: Positive Feedback Regulation of Type I IFN Production by the IFN-Inducible DNA Sensor cGAS
      • Article
      • Peer Reviewed
      UCLA Previously Published Works (2014)
      The retinoid X receptor α (RXRα), a key nuclear receptor in metabolic processes, is downregulated during host antiviral response. However, the roles of RXRα in host antiviral response are unknown. Here we show that RXRα overexpression or ligand activation increases host susceptibility to viral infections in vitro and in vivo, while Rxra-/- or antagonist treatment reduces infection by the same viruses. Consistent with these functional studies, ligand activation of RXR inhibits the expression of antiviral genes including type I interferon (IFN) and Rxra-/- macrophages produce more IFNβ than WT macrophages in response to polyI:C stimulation. Further results indicate that ligand activation of RXR suppresses the nuclear translocation of β-catenin, a co-activator of IFNβ enhanceosome. Thus, our studies have uncovered a novel RXR-dependent innate immune regulatory pathway, suggesting that the downregulation of RXR expression or RXR antagonist treatment benefits host antiviral response, whereas RXR agonist treatment may increase the risk of viral infections.
        Cover page: Retinoid X receptor α attenuates host antiviral response by suppressing type I interferon
        • Article
        • Peer Reviewed
        UCLA Previously Published Works (2015)
        The candidate phylum TM7 is globally distributed and often associated with human inflammatory mucosal diseases. Despite its prevalence, the TM7 phylum remains recalcitrant to cultivation, making it one of the most enigmatic phyla known. In this study, we cultivated a TM7 phylotype (TM7x) from the human oral cavity. This extremely small coccus (200-300 nm) has a distinctive lifestyle not previously observed in human-associated microbes. It is an obligate epibiont of an Actinomyces odontolyticus strain (XH001) yet also has a parasitic phase, thereby killing its host. This first completed genome (705 kb) for a human-associated TM7 phylotype revealed a complete lack of amino acid biosynthetic capacity. Comparative genomics analyses with uncultivated environmental TM7 assemblies show remarkable conserved gene synteny and only minimal gene loss/gain that may have occurred as TM7x adapted to conditions within the human host. Transcriptomic and metabolomic profiles provided the first indications, to our knowledge, that there is signaling interaction between TM7x and XH001. Furthermore, the induction of TNF-α production in macrophages by XH001 was repressed in the presence of TM7x, suggesting its potential immune suppression ability. Overall, our data provide intriguing insights into the uncultivability, pathogenicity, and unique lifestyle of this previously uncharacterized oral TM7 phylotype.
          Cover page: Cultivation of a human-associated TM7 phylotype reveals a reduced genome and epibiotic parasitic lifestyle.
          • Article
          • Peer Reviewed
          UC San Francisco Previously Published Works (2021)
          Diet is a major factor that shapes the gut microbiome1, but the consequences of diet-induced changes in the microbiome for host pathophysiology remain poorly understood. We conducted a randomized human intervention study using a very-low-calorie diet (NCT01105143). Although metabolic health was improved, severe calorie restriction led to a decrease in bacterial abundance and restructuring of the gut microbiome. Transplantation of post-diet microbiota to mice decreased their body weight and adiposity relative to mice that received pre-diet microbiota. Weight loss was associated with impaired nutrient absorption and enrichment in Clostridioides difficile, which was consistent with a decrease in bile acids and was sufficient to replicate metabolic phenotypes in mice in a toxin-dependent manner. These results emphasize the importance of diet-microbiome interactions in modulating host energy balance and the need to understand the role of diet in the interplay between pathogenic and beneficial symbionts.
            Cover page: Caloric restriction disrupts the microbiota and colonization resistance
            • Article
            • Peer Reviewed
            UCLA Previously Published Works (2017)
            New influenza vaccines that provide effective and broad protection are desperately needed. Live attenuated viruses are attractive vaccine candidates because they can elicit both humoral and cellular immune responses. However, recent formulations of live attenuated influenza vaccines (LAIVs) have not been protective. We combined high-coverage transposon mutagenesis of influenza virus with a rapid high-throughput screening for attenuation to generate W7-791, a live attenuated mutant virus strain. W7-791 produced only a transient asymptomatic infection in adult and neonatal mice even at doses 100-fold higher than the LD50 of the parent strain. A single administration of W7-791 conferred full protection to mice against lethal challenge with H1N1, H3N2, and H5N1 strains, and improved viral clearance in ferrets. Adoptive transfer of T cells from W7-791-immunized mice conferred heterologous protection, indicating a role for T cell-mediated immunity. These studies present an LAIV development strategy to rapidly generate and screen entire libraries of viral clones.
              Cover page: Generation of a Live Attenuated Influenza Vaccine that Elicits Broad Protection in Mice and Ferrets
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