嗜神经病毒躲避宿主先天性免疫反应的机制和应用培训课件

1、嗜神经病毒躲避宿主先天性免疫反应的机制和应用嗜神经病毒躲避宿主先天性免疫反应的机制和应用Rabies virus180nm x 75nm嗜神经病毒躲避宿主先天性免疫反应的机制和应用2Rabies virus180nm x 75nm嗜神经病毒躲Robert Hurt-USCRabies pathogenesisPatients die of circulatory insufficiency, cardiac arrest and respiratory failure.嗜神经病毒躲避宿主先天性免疫反应的机制和应用3Robert Hurt-USCRabies pathogen4Rabies in

2、fection and innate immunityWang et al. Journal of Virology, 2005嗜神经病毒躲避宿主先天性免疫反应的机制和应用44Rabies infection and innate i5Zhao et al. Journal of Virology, 2009重组狂犬病病毒构建嗜神经病毒躲避宿主先天性免疫反应的机制和应用55Zhao et al. Journal of Virolo6Expression of MIP-1 attenuated RABV pathogenicity, while expression of RANTES or I

3、P-10 increased RABV pathogencityZhao et al. Journal of Virology, 2009嗜神经病毒躲避宿主先天性免疫反应的机制和应用66Expression of MIP-1 attenuat7Control rHEP HEP-MIP1a HEP-RANTES HEP-IP10D3D6D9 HE staining of mouse brainsZhao et al. Journal of Virology, 2009嗜神经病毒躲避宿主先天性免疫反应的机制和应用77Control rHEP Expression of MIP-1 enhances

4、 VNA production and protectionZhao et al. Journal of Virology, 2010嗜神经病毒躲避宿主先天性免疫反应的机制和应用8Expression of MIP-1 enhancesZhao et al. Journal of Virology, 2010外周过量表达MIP-1会吸引更多的树突状细胞和B细胞嗜神经病毒躲避宿主先天性免疫反应的机制和应用9Zhao et al. Journal of Virolog嗜神经病毒躲避宿主先天性免疫反应的机制和应用10嗜神经病毒躲避宿主先天性免疫反应的机制和应用10 Antigenic GroupVi

5、rusHostDiseaseI229ENL63humanhumanrespiratory infectionrespiratory infection, croupTGEVpigrespiratory and enteric infectionCCVdogenteric infectionFECVcatenteric infectionFIPVcatrespiratory,enteric, hepatitis and neurological infectionIIOC43humanRespiratory and possibly enteric infectionSARS-CoVHKU1hu

6、manhumanRespiratoryRespiratoryMHVmouseRespiratory, enteric, neurologic infectionHEVpigRespiratory, enteric, neurologic infectionBCVcowEnteric infectionTCVturkeyRespiratory and enteric infection IIIIBVchickenRespiratory and enteric infection, hepatitisCoronaviruses cause diseases in humans and domest

7、ic animalsAdapted from Holmes and Lai, Fields Virology嗜神经病毒躲避宿主先天性免疫反应的机制和应用11 Antigenic GroupVirusHostDiseaWhy MHV?MHV produces a broad spectrum of disease in the mouse -pneumonia (MHV-1) -hepatitis (MHV-A59) -encephalitis (MHV-A59/JHM) -demylination (MHV-A59)It provides excellent small animal mode

8、ls for hepatitis, for SARS, and for multiple sclerosis嗜神经病毒躲避宿主先天性免疫反应的机制和应用12Why MHV?MHV produces a broad sPart I: MHV ns2 interferes type I interferon responses 嗜神经病毒躲避宿主先天性免疫反应的机制和应用13Part I: MHV ns2 interferes tMutation of ns2 confers attenuation of hepatitis but not CNS disease IC 500PFUIH 500P

9、FU Roth-Cross, J.K. et al, JVI, 2009, 83(8):3743-3753.brainliver嗜神经病毒躲避宿主先天性免疫反应的机制和应用14Mutation of ns2 confers attenNs2 is an organ specific virulence factor 1a1b2a45a5bHESEMINMHV 2H phosphodiesterase; 2 predicted catalytic His-x-Thr/Ser motifs (Mazumder et a., 2002; Snijder et al.,2003)ns2(Mazumbd

10、er et al., 2002Snijder et al.,2003; Roth-Cross, 2009)嗜神经病毒躲避宿主先天性免疫反应的机制和应用15Ns2 is an organ specific virulMutation of ns2 confers attenuation of replication in macrophages and microglia but not in other cell types Zhao, L. et al, JVI, 2011.Oct; 85(19):10058-10068. 嗜神经病毒躲避宿主先天性免疫反应的机制和应用16Mutation o

11、f ns2 confers attenuMOI 1MOI 0.01 ns2 mutants recover the ability to replicate efficiently in macrophages and microglia from IFNAR knockout miceMOI 1 Zhao, L. et al, JVI, 2011.Oct; 85(19):10058-10068. 嗜神经病毒躲避宿主先天性免疫反应的机制和应用17MOI 1MOI 0.01 ns2 mutants recoType 1 interferon induction and signaling pat

12、hwaysIFN-IFNTYK2JAK1PSTAT1P STAT1 orSTAT2IFNAR1IFNAR2ISREIRF9PSTAT1PSTAT2 ISGF3PSTAT2IRF-9ISGs:OAS, MxA, ISG15, ISG54,MDA5TLR3TLR7/8TLR9dsRNACpGssRNATIRTBK-1IKK-IRF-3IFN-NF-BATF-2RIG-ICBPPMDA5TIRTIRTRIFMyD88MyD88ISREIFN-嗜神经病毒躲避宿主先天性免疫反应的机制和应用18Type 1 interferon induction anL2AstroBMMIFNAR-/- BMMns2

13、mutants are not defective in induction of IFN-/ mRNABoth wt RA59 and ns2 mutants induce minimal amounts of IFN-, mRNA in L2 cells and astrocytes wt RA59 and ns2 mutants induce similar levels of IFN-, mRNA in BMM from both B6 and IFNAR-/- mice Zhao, L. et al, JVI, 2011.Oct; 85(19):10058-10068. 嗜神经病毒躲

14、避宿主先天性免疫反应的机制和应用19L2AstroBMMIFNAR-/- BMMns2 mutaBMMBMMMicroglians2 mutants are more sensitive to the antiviral effects of IFN-/ than wt A59 in macrophages and microgliabut not in other cell typesL2Astro嗜神经病毒躲避宿主先天性免疫反应的机制和应用20BMMBMMMicroglians2 mutants areViral replication and IFN sensitivity in the

15、 hepatocytesNo IFNIFN嗜神经病毒躲避宿主先天性免疫反应的机制和应用21Viral replication and IFN sensIn vivo macrophage depletionLiposomes, encapsulating the Clodronate molecules (squares), are ingested by macrophages via endocytosis. After fusion with lysosomes (L) containing phospholipases (arrowheads), the latter disrupt

16、the bilayers of the liposomes. The more concentric bilayers are disrupted, the greater is the Clodronate release within the cell. The cells are killed by Clodronate through apoptosis. 嗜神经病毒躲避宿主先天性免疫反应的机制和应用22In vivo macrophage depletionLiLiposome ClodronateA59ns2-H126RA59ns2-H126RPBSLiposome Clodron

17、ateH&E stainingN protein stainingns2 mutants replicate and induce hepatitis in macrophage depleted mice500 fold vs 10 fold嗜神经病毒躲避宿主先天性免疫反应的机制和应用23Liposome ClodronateA59ns2-H126嗜神经病毒躲避宿主先天性免疫反应的机制和应用培训课件Part I: conclusionsns2 is an organ specific virulence factor and antagonizes IFN signalingns2 is r

18、equired for replication in macrophages; depletion of macrophages in vivo promotes ns2 mutant virus replicationwe suggest that MHV has to replicate in Kupffer cells in the liver sinusoids in order to reach the liver parenchyma and induce hepatitis 嗜神经病毒躲避宿主先天性免疫反应的机制和应用25Part I: conclusions嗜神经病毒躲避宿主先

19、天Type 1 interferon induction and signaling pathwaysIFN-IFNTYK2JAK1PSTAT1P STAT1 orSTAT2IFNAR1IFNAR2ISREIRF9PSTAT1PSTAT2 ISGF3PSTAT2IRF-9ISGs:OAS, MxA, ISG15, ISG54,MDA5TLR3TLR7/8TLR9dsRNACpGssRNATIRTBK-1IKK-IRF-3IFN-NF-BATF-2RIG-ICBPPMDA5TIRTIRTRIFMyD88MyD88ISREIFN-嗜神经病毒躲避宿主先天性免疫反应的机制和应用26Type 1 int

20、erferon induction anpCAGGS-IFNNS2a-IFNSV5V-IFNNDV-bioassay in Vero cellspCAGGS-no IFNTansfection of pCAGGSor pCAGGS-ns2 or pCAGGS-SV5Vin Vero cellsTreatment with or w/o1000U/ml for 16hInfection of NDV-GFP24h12hGFP 嗜神经病毒躲避宿主先天性免疫反应的机制和应用27pCAGGS-IFNNS2a-IFNSV5V-IFNNDV-ISG screening in KO BMMISG15IFIT

21、1IFIT2PKRRNase L嗜神经病毒躲避宿主先天性免疫反应的机制和应用28ISG screening in KO BMMISG15IFPart II: MHV ns2 antagonizes OAS-RNase L pathway嗜神经病毒躲避宿主先天性免疫反应的机制和应用29Part II: MHV ns2 antagonizesInterferon signaling modelViral dsRNAMDA5*, RIG-I*IFNAntiviral ISGsOAS*2-5ARNase LCellular and viral RNASmall RNAs2-PDEns2?Main pa

22、thwayOAS-RNase L pathwayOAS= 2,5-oliogoadenylate synthetase2-PDE= 2 phosphodiesterase2-5A=2,5oligoadenylate嗜神经病毒躲避宿主先天性免疫反应的机制和应用30Interferon signaling modelViraWild type A59 replication in BMM was not affected by the OAS-RNase L system嗜神经病毒躲避宿主先天性免疫反应的机制和应用31Wild type A59 replication in BDefective

23、replication of ns2 mutant is restored in RNase L-/- BMMBut not in PKR-/- BMMZhao, L et al. Cell Host & Microbe, 2012, (11) 607616.嗜神经病毒躲避宿主先天性免疫反应的机制和应用32Defective replication of ns2 mns2 expression in 293T cellspCAGGS-ns2pCAGGSpCAGGS-ns2-H126RZhao, L et al. Cell Host & Microbe, 2012, (11) 607616.嗜神

24、经病毒躲避宿主先天性免疫反应的机制和应用33ns2 expression in 293T cellspCns2 prevents rRNA cleavage and 2-5A production in BMMZhao, L et al. Cell Host & Microbe, 2012, (11) 607616.嗜神经病毒躲避宿主先天性免疫反应的机制和应用34ns2 prevents rRNA cleavage andOverexpression of ns2 in 293T cells prevents rRNA cleavage and 2-5A production induced by poly I:CZhao, L et al. Cell Host & Microbe, 2012, (11) 607616.嗜神经病毒躲避宿主先天性免疫反应的机制和应用35Overexpression of ns2 in 293T Ns2 cleaves 2-5A into ATP and AMPZhao, L et al. Cell Host & Microbe, 2012, (11) 607616.嗜神经病毒躲避宿主先天性免疫反应的机制和应用36Ns2 cleaves 2-5A into ATP and Ns2 enhances