The proteome of Toll-like receptor 3–stimulated human immortalized fibroblasts: Implications for susceptibility to herpes simplex virus encephalitis

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Presentation transcript:

The proteome of Toll-like receptor 3–stimulated human immortalized fibroblasts: Implications for susceptibility to herpes simplex virus encephalitis Rebeca Pérez de Diego, PhD, Claire Mulvey, PhD, Mark Crawford, BSc, Matthew W.B. Trotter, PhD, Lazaro Lorenzo, BSc, Vanessa Sancho-Shimizu, PhD, Laurent Abel, MD, PhD, Shen-Ying Zhang, MD, PhD, Jean-Laurent Casanova, MD, PhD, Jasminka Godovac- Zimmermann, PhD Journal of Allergy and Clinical Immunology Volume 131, Issue 4, Pages 1157-1166 (April 2013) DOI: 10.1016/j.jaci.2013.01.008 Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 1 A, Illustration of the SILAC workflow. B, Summary of samples used and TLR3 pathway state for the SILAC experiment. C, Overview of MS results obtained with MaxQuant software. C1-3, Healthy controls; C2NS, healthy control-2 nonstimulated; H, heavy medium; L, light medium; NS, nonstimulated cells; P, patient with unknown gene defect; TLR3, poly(I:C)-stimulated cells; UNC-93B−/−, UNC-93B–deficient patient; WT, wild-type. *Protein groups identified with protein and peptide false discovery rate = 1%. †Protein abundance ratios calculated as median values using a minimum of 3 quantifiable razor peptides. ‡Protein ratios with significance B ≤.05 and SILAC ratios ≥1.5. Journal of Allergy and Clinical Immunology 2013 131, 1157-1166DOI: (10.1016/j.jaci.2013.01.008) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 2 Production of IFN-β by SV40 fibroblasts after poly(I:C) stimulation for 24 hours as assessed by ELISA. Mean values ± SD were calculated from 3 independent experiments. C1-C5, Positive healthy controls; UNC93B1−/−, UNC-93B–deficient patient. Journal of Allergy and Clinical Immunology 2013 131, 1157-1166DOI: (10.1016/j.jaci.2013.01.008) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 3 A, For proteins with potential biological significance in HSE, the SILAC ratio detected for healthy controls (C1-3), healthy control 2 nonstimulated (C2NS), UNC-93B–deficient patient (UNC-93B−/−), and patient with unknown gene defect (P). Significance B P value <.05. B, Illustration of potential biological significance in immunity against HSE for proteins upregulated after TLR3 activation. ds, Double stranded. Journal of Allergy and Clinical Immunology 2013 131, 1157-1166DOI: (10.1016/j.jaci.2013.01.008) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 4 Human SV40 fibroblast nontransfected (−), transfected with SOD2 or Mock vector. A, Western-blot analysis of SOD2 levels. Densitometry normalized with respect to glyceraldehyde-3-phosphate dehydrogenase, expressed as a fold induction over nonstimulated/nontransfected control cells. The panel is representative of 3 experiments. B, Caspase-3/7 activity in nonstimulated (NS) or poly(I:C)-stimulated cells. Luciferase activity represented as percentage; 100% is healthy control nonstimulated. Mean values ± SD were calculated from 3 independent experiments. Two-sided test was used, and a P value of less than .05 was considered statistically significant. C, Healthy control C3; UNC-93B−/−, UNC-93B–deficient patient; −, nonstimulated; +, poly(I:C) stimulated. Journal of Allergy and Clinical Immunology 2013 131, 1157-1166DOI: (10.1016/j.jaci.2013.01.008) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig E1 Journal of Allergy and Clinical Immunology 2013 131, 1157-1166DOI: (10.1016/j.jaci.2013.01.008) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig E2 Journal of Allergy and Clinical Immunology 2013 131, 1157-1166DOI: (10.1016/j.jaci.2013.01.008) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig E3 Journal of Allergy and Clinical Immunology 2013 131, 1157-1166DOI: (10.1016/j.jaci.2013.01.008) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig E4 Journal of Allergy and Clinical Immunology 2013 131, 1157-1166DOI: (10.1016/j.jaci.2013.01.008) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig E5 Journal of Allergy and Clinical Immunology 2013 131, 1157-1166DOI: (10.1016/j.jaci.2013.01.008) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig E6 Journal of Allergy and Clinical Immunology 2013 131, 1157-1166DOI: (10.1016/j.jaci.2013.01.008) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions