28 research outputs found

    Homozygous NLRP1 gain-of-function mutation in siblings with a syndromic form of recurrent respiratory papillomatosis

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    © 2019 National Academy of Sciences. All rights reserved. Juvenile-onset recurrent respiratory papillomatosis (JRRP) is a rare and debilitating childhood disease that presents with recurrent growth of papillomas in the upper airway. Two common human papillomaviruses (HPVs), HPV-6 and -11, are implicated in most cases, but it is still not understood why only a small proportion of children develop JRRP following exposure to these common viruses. We report 2 siblings with a syndromic form of JRRP associated with mild dermatologic abnormalities. Whole-exome sequencing of the patients revealed a private homozygous mutation in NLRP1, encoding Nucleotide-Binding Domain Leucine-Rich Repeat Family Pyrin Domain-Containing 1. We find the NLRP1 mutant allele to be gain of function (GOF) for inflammasome activation, as demonstrated by the induction of inflammasome complex oligomerization and IL-1ÎČ secretion in an overexpression system. Moreover, patient-derived keratinocytes secrete elevated levels of IL-1ÎČ at baseline. Finally, both patients displayed elevated levels of inflammasome-induced cytokines in the serum. Six NLRP1 GOF mutations have previously been described to underlie 3 allelic Mendelian diseases with differing phenotypes and modes of inheritance. Our results demonstrate that an autosomal recessive, syndromic form of JRRP can be associated with an NLRP1 GOF mutation

    Cd81 Interacts with the T Cell Receptor to Suppress Signaling

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    CD81 (TAPA-1) is a ubiquitously expressed tetraspanin protein identified as a component of the B lymphocyte receptor (BCR) and as a receptor for the Hepatitis C Virus. In an effort to identify trans-membrane proteins that interact with the T-cell antigen receptor (TCR), we performed a membrane yeast two hybrid screen and identified CD81 as an interactor of the CD3delta subunit of the TCR. We found that in the absence of CD81, in thymocytes from knockout mice, TCR engagement resulted in stronger signals. These results were recapitulated in T cell lines that express low levels of CD81 through shRNA mediated silencing. Increased signaling did not result from alterations in the levels of TCR on the surface of T lymphocytes. Although CD81 is not essential for normal T lymphocyte development, it plays an important role in regulating TCR and possibly pre-TCR signal transduction by controlling the strength of signaling. CD81 dependent alterations in thymocyte signaling are evident in increased CD5 expression on CD81 deficient double positive (DP) thymocytes. We conclude that CD81 interacts with the T cell receptor to suppress signaling. © 2012 Cevik et al

    Dynamic Modulation of Thymic MicroRNAs in Response to Stress

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    thymocyte subsets. Several of the differentially regulated murine thymic miRs are also stress responsive in the heart, kidney, liver, brain, and/or spleen. The most dramatic thymic microRNA down modulated is miR-181d, exhibiting a 15-fold reduction following stress. This miR has both similar and distinct gene targets as miR-181a, another member of miR-181 family. Many of the differentially regulated microRNAs have known functions in thymopoiesis, indicating that their dysregulation will alter T cell repertoire selection and the formation of naĂŻve T cells. This data has implications for clinical treatments involving anti-inflammatory steroids, ablation therapies, and provides mechanistic insights into the consequences of infections

    Life-threatening influenza pneumonitis in a child with inherited IRF9 deficiency

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    Life-threatening pulmonary influenza can be caused by inborn errors of type I and III IFN immunity. We report a 5-yr-old child with severe pulmonary influenza at 2 yr. She is homozygous for a loss-of-function IRF9 allele. Her cells activate gamma-activated factor (GAF) STAT1 homodimers but not IFN-stimulated gene factor 3 (ISGF3) trimers (STAT1/STAT2/IRF9) in response to IFN-α2b. The transcriptome induced by IFN-α2b in the patient's cells is much narrower than that of control cells; however, induction of a subset of IFN-stimulated gene transcripts remains detectable. In vitro, the patient's cells do not control three respiratory viruses, influenza A virus (IAV), parainfluenza virus (PIV), and respiratory syncytial virus (RSV). These phenotypes are rescued by wild-type IRF9, whereas silencing IRF9 expression in control cells increases viral replication. However, the child has controlled various common viruses in vivo, including respiratory viruses other than IAV. Our findings show that human IRF9- and ISGF3-dependent type I and III IFN responsive pathways are essential for controlling IAV

    Inborn errors of OAS-RNase L in SARS-CoV-2-related multisystem inflammatory syndrome in children

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    Multisystem inflammatory syndrome in children (MIS-C) is a rare and severe condition that follows benign COVID-19. We report autosomal recessive deficiencies of OAS1, OAS2, or RNASEL in five unrelated children with MIS-C. The cytosolic double-stranded RNA (dsRNA)-sensing OAS1 and OAS2 generate 2'-5'-linked oligoadenylates (2-5A) that activate the single-stranded RNA-degrading ribonuclease L (RNase L). Monocytic cell lines and primary myeloid cells with OAS1, OAS2, or RNase L deficiencies produce excessive amounts of inflammatory cytokines upon dsRNA or severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) stimulation. Exogenous 2-5A suppresses cytokine production in OAS1-deficient but not RNase L-deficient cells. Cytokine production in RNase L-deficient cells is impaired by MDA5 or RIG-I deficiency and abolished by mitochondrial antiviral-signaling protein (MAVS) deficiency. Recessive OAS-RNase L deficiencies in these patients unleash the production of SARS-CoV-2-triggered, MAVS-mediated inflammatory cytokines by mononuclear phagocytes, thereby underlying MIS-C

    Genetic Basis of Myocarditis: Myth or Reality?

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    Transgenic Expression of MicroRNA-181d Augments the Stress-Sensitivity of CD4<sup>+</sup>CD8<sup>+</sup> Thymocytes

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    <div><p>Physiological stress resulting from infections, trauma, surgery, alcoholism, malnutrition, and/or pregnancy results in a substantial depletion of immature CD4<sup>+</sup>CD8<sup>+</sup> thymocytes. We previously identified 18 distinct stress-responsive microRNAs (miRs) in the thymus upon systemic stress induced by lipopolysaccharide (LPS) or the synthetic glucocorticoid, dexamethasone (Dex). MiRs are short, non-coding RNAs that play critical roles in the immune system by targeting diverse mRNAs, suggesting that their modulation in the thymus in response to stress could impact thymopoiesis. MiR-181d is one such stress-responsive miR, exhibiting a 15-fold down-regulation in expression. We utilized both transgenic and gene-targeting approaches to study the impact of miR-181d on thymopoiesis under normal and stress conditions. The over-expression of miR-181d in developing thymocytes reduced the total number of immature CD4<sup>+</sup>CD8<sup>+</sup> thymocytes. LPS or Dex injections caused a 4-fold greater loss of these cells when compared with the wild type controls. A knockout mouse was developed to selectively eliminate miR-181d, leaving the closely spaced and contiguous family member miR-181c intact. The targeted elimination of just miR-181d resulted in a thymus stress-responsiveness similar to wild-type mice. These experiments suggest that one or more of three other miR-181 family members have overlapping or compensatory functions. Gene expression comparisons of thymocytes from the wild type versus transgenic mice indicated that miR-181d targets a number of stress, metabolic, and signaling pathways. These findings demonstrate that selected miRs enhance stress-mediated thymic involution <i>in vivo</i>.</p></div

    MiR-181d transgenic mice.

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    <p>(A) Schematic shows the sequence homology between mature miR-181 family members. 5â€Č-seed region is underlined. Base differences are shaded with gray. (B) MiR-181d expression in various tissues examined by Northern blotting. U6 probe was used as the endogenous control. (C) Cloning of the pri-miR-181d into the VA-hCD2 transgenic cassette. Stem-loop structure of pre-miR-181d is shown, in which mature miR-181d is highlighted in blue. (D) Relative miR-181d levels were determined by real-time quantitative PCR. Littermate control values were set to 1. Graph represents the mean fold changes +/− SEM normalized to the U6 levels from 3 independent samples, performed in triplicates (n.s.  =  non-significant, *<i>p</i><0.05, **<i>p</i><0.01, ***<i>p</i><0.001; Two-tailed unpaired Student's <i>t</i>-test).</p

    MiR-181d over-expression reduces the number of DP thymocytes.

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    <p>(A) Total thymus cellularity in the control and miR-181d Tg mice. (B) Representative plots show CD4 by CD8 profiles of thymocytes in the control and miR-181d Tg mice, analyzed by FACS. (C) Average percentages of thymocyte subsets (DN, DP, CD4 SP, and CD8 SP) from the control and miR-181d Tg mice. (D) Absolute cell numbers of DP thymocytes. (E) Absolute cell numbers of CD4 SP (left) and CD8 SP (right) thymocytes. (A–E) Data are from WT (n = 18), Tg-8 (n = 25), and Tg-38 (n = 16) mice. (F) Total thymus cellularity of the OTII Tg and OTII/miR-181d Tg-38 mice. (G) Total thymocytes were stained for CD4 and CD8, and analyzed by FACS. (H) Average percentages of DP and CD4 SP thymocytes are shown. (I) Histogram shows the surface expression of TCR (TCR Vα2) gated on CD4<sup>+</sup>CD8<sup>−</sup> SP thymocytes from the OTII Tg (dark gray) and OTII/miR-181d Tg-38 mice (black line). (F–I) Data are from at least 2 mice per group. Each bar is the mean +/− SEM (n.s.  =  non-significant, *<i>p</i><0.05, **<i>p</i><0.01, ***<i>p</i><0.001; Two-tailed unpaired Student's <i>t</i>-test). (J) Histograms show CD69 expression on CD4 SP and CD8 SP thymocytes from the WT (white), Tg-8 (light gray), and Tg-38 (dark gray) mice. (K) Relative MFI (Mean Fluorescence Intensity) levels of CD69 on SP thymocytes. (L) Ratio of the CD69<sup>+</sup>TCRÎČ<sup>high</sup> to CD69<sup>−</sup>TCRÎČ<sup>high</sup> thymocyte numbers shown for CD4 SP and CD8 SP thymocytes. (M) Average percentages of Annexin V<sup>+</sup> cells gated on DP thymocytes. (J-M) Data are of at least 3 mice per group. All bar graphs represent the mean +/− SEM values (n.s.  =  non-significant, *<i>p</i><0.05, **<i>p</i><0.01, ***<i>p</i><0.001; One-way ANOVA followed by Tukey's post-hoc test).</p
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