Biology subjects

Ko, N.

Publications and source records attributed to Ko, N..

4 recordsLinked to original sources

bioRxiv2024

The loss of both pUL16 and pUL21 in HSV-1 infected cells abolishes cytoplasmic envelopment.

Previously, we had developed synthetic genomics methods to assemble an infectious clone of herpes simplex virus type-1 (HSV-1). To do this, the genome was assembled from 11 separate cloned fragments in yeast using transformation associated recombination. The eleven fragments or "parts" spanned the 152 kb genome and recombination was achieved because of the overlapping homologous sequences between each fragment. To demonstrate the robustness of this genome assembly method for reverse genetics, we engineered different mutations that were located in distant loci on the genome and built a collection of HSV-1 genomes that contained single and different combination of mutations in 5 conserved HSV-1 genes. The five genes: UL7, UL11, UL16, UL21 and UL51 encode virion structural proteins and have varied functions in the infected cell. Each is dispensable for virus replication in cell culture, however, combinatorial analysis of deletions in the five genes revealed "synthetic-lethality" of some of the genetic mutations. Thus, it was discovered that any virus that carried a UL21 mutation in addition to the other gene was unable to replicate in Vero cells. Replication was restored in a complementing cell line that provided pUL21 in trans. One particular combination (UL16-UL21) was of interest because the proteins encoded by these genes are known to physically interact and are constituents of the tegument structure. Furthermore, their roles in HSV-1 infected cells are unclear. Both are dispensable for HSV-1 replication, however, in HSV-2 their mutation results in nuclear retention of assembled capsids. We thus characterized these viruses that carry the single and double mutant. What we discovered is that in cells where both pUL16 and pUL21 are absent, cytoplasmic capsids were evident but did not mature into enveloped particles. The capsid particles isolated from these cells showed significantly lower levels of incorporation of both VP16 and pUL37 when compared to the wild-type capsids. These data now show that of the tegument proteins, like the essential pUL36, pUL37 and VP16; the complex of pUL16 and pUL21 should be considered as important mediators of cytoplasmic maturation of the particle.

microbiology↗

bioRxiv2020

Autocrine IL11 cis-signaling in hepatocytes is an initiating nexus between lipotoxicity and non-alcoholic steatohepatitis

Background and aimsIL11 signaling is important in non-alcoholic steatohepatitis (NASH) but how it contributes to NASH pathologies beyond fibrosis is not known. Here we investigate the role of IL11 signaling in hepatocyte lipotoxicity. MethodsHepatocytes were stimulated with IL6, IL11, HyperIL6, or HyperIL11 alone or in the presence of soluble gp130 (sgp130) or soluble IL11RA (sIL11RA), or loaded with palmitate in the presence of IgG or anti-IL11RA (X209) antibodies or sgp130. Effects were assessed using colorimetric ALT, GSH, or ELISA assays, immunoblots, and flow cytometry. The relative contributions of IL11 cis-versus -trans signaling in vivo was assessed in two preclinical NASH models using a high fat methionine/choline deficient diet or a Western diet with liquid fructose in C57BL6/Ntac mice injected with AAV8-Alb-Cre, AAV8-Alb-sgp130, in mice with hepatocyte-specific deletion of Il11ra (CKO), and in mice with global deletion of Il11ra injected with AAV8-Alb-mIl11ra or AAV8-Alb-sIl11ra. Livers and serum were collected; serum samples were analyzed using biochemistry and liver tissues were analyzed by histology, qPCR, immunobloting, hydroxyproline, and GSH assays. ResultsWe show that lipid-laden hepatocytes secrete IL11, which acts via autocrine cis-signaling to cause lipoapoptosis. IL11 causes lipotoxic hepatocyte death through activation of non-canonical signaling pathways and increased NOX4-derived reactive oxygen species. In two preclinical models, hepatocyte-specific deletion of Il11ra1 protects mice from all aspects of NASH with beneficial effects on body weight. In accordance, restoration of IL11 cis-signaling in hepatocytes only in mice globally deleted for Il11ra1 reconstitutes steatosis and inflammation. Throughout, we found no evidence to support the existence of IL6 or IL11 trans-signaling in the liver. ConclusionWe conclude that autocrine IL11-mediated cell death underlies hepatocyte lipotoxicity and that liver fibrosis and inflammation occur subsequently. These data highlight a new disease mechanism for the transition from compensated fatty liver disease to NASH.

molecular biology↗

bioRxiv2019

Interleukin 11 expression causes murine inflammatory bowel disease

Interleukin 11 (IL11) is a profibrotic cytokine, secreted by myofibroblasts and damaged epithelial cells. Smooth muscle cells (SMCs) also secrete IL11 under pathological conditions and express the IL11 receptor. Here we examined the effects of SMC-specific, conditional expression of murine IL11 in a transgenic mouse (Il11SMC). Within days of transgene activation, Il11SMC mice developed loose stools and progressive bleeding and rectal prolapse, which was associated with a 65% mortality by two weeks. The bowel of Il11SMC mice was inflamed, fibrotic and had a thickened wall, which was accompanied by activation of ERK and STAT3. In other organs, including heart, lung, liver, kidney and skin there was a phenotypic spectrum of fibro-inflammation, together with consistent ERK activation. To investigate further the importance of stromal-derived IL11 in the inflammatory bowel phenotype we used a second model with fibroblast-specific expression of IL11, the Il11Fib mouse. This additional model largely phenocopied the Il11SMC bowel phenotype. These data show that IL11 secretion from the stromal niche is sufficient to drive inflammatory bowel disease in mice. Given that IL11 expression in colonic stromal cells predicts anti-TNF therapy failure in patients with ulcerative colitis or Crohns disease, we suggest IL11 as a therapeutic target for inflammatory bowel disease.

pathology↗

bioRxiv2018

IL-11 is a therapeutic target in idiopathic pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) remains a progressive disease despite best medical management. We previously identified IL-11 as a critical factor for cardiovascular fibrosis and examine here its role in pulmonary fibrosis. IL-11 is consistently upregulated in IPF genomic datasets, which we confirmed by histology. Pulmonary fibroblasts stimulated with IL-11 transform into invasive myofibroblasts whereas fibroblasts from Il11ra deleted mice did not respond to pro-fibrotic stimuli. In the mouse, injection of recombinant Il-11 or fibroblast-specific expression of Il-11 caused pulmonary fibrosis. We then generated a neutralising IL-11 binding antibody that blocks lung fibroblast activation across species. In a mouse model of IPF, anti-IL-11 therapy attenuated lung fibrosis and specifically blocked Erk activation. These data prioritise IL-11 as an accessible drug target in IPF.\n\nOne Sentence SummaryNon-canonical IL-11 signalling is a central hallmark of idiopathic pulmonary fibrosis and represents a novel target for antibody therapies.

molecular biology↗