Date post: | 29-Dec-2015 |
Category: |
Documents |
Upload: | osborne-gardner |
View: | 214 times |
Download: | 0 times |
Response of the Innate Response of the Innate Immune System to Immune System to
Pathogens:Pathogens:Pattern Recognition Pattern Recognition
ReceptorsReceptors
What is the Innate Immune What is the Innate Immune Response?Response?
A universal and evolutionarily conserved mechanism A universal and evolutionarily conserved mechanism of host defense against infectionof host defense against infection
First line of DefenseFirst line of Defense Predates the adaptive immune responsePredates the adaptive immune response
Found in all multicellular organismsFound in all multicellular organisms Adaptive only in vertebratesAdaptive only in vertebrates
Uses receptors and effectors that are ancient in Uses receptors and effectors that are ancient in their lineagetheir lineage
Must provide protection against a wide variety of Must provide protection against a wide variety of pathogenspathogens
Distinguishes self from non-self perfectlyDistinguishes self from non-self perfectly Defects in innate immunity are very rare and almost Defects in innate immunity are very rare and almost
always lethalalways lethal
The Innate Immune Response:The Innate Immune Response:Common MisconceptionsCommon Misconceptions
The innate immune system is an evolutionary rudiment The innate immune system is an evolutionary rudiment whose only function is to contain the infection until the whose only function is to contain the infection until the “real” immune response can kick in.“real” immune response can kick in.
Adaptive immunity developed because of the inflexibility Adaptive immunity developed because of the inflexibility of the nonclonal receptors used by the innate immune of the nonclonal receptors used by the innate immune response. The innate system cannot cope with the high response. The innate system cannot cope with the high mutational rate and heterogeneity of pathogenic mutational rate and heterogeneity of pathogenic organisms.organisms.
The Innate immune system instructs the adaptive The Innate immune system instructs the adaptive immune response to respond to microbial infectionimmune response to respond to microbial infection
The major decision to respond or not respond to The major decision to respond or not respond to a particular ligand is decided by the genome-a particular ligand is decided by the genome-
encoded receptors of the innate immune systemencoded receptors of the innate immune system
Y
Endosome
YNaive T Cell
MHC B7
Inflammatory and effectorcytokines
PRR
PAMP
Activated T Cell
CD40L, FasL, CD30L, CD27L
B Cell
Adapted from Medzhitov and JanewayCur. Opin. Immunol. 1997 9:4-9
Phagocytosis
APC
Direct Bactericidal ActivityPhagocytosisOxygen burstAnti-microbial peptides
Pathogen-specific Antibody
Complement
Innate Immunity Adaptive Immunity
skin, gut villi, lung cilia,etc
many protein andnon-protein secretions
phagocytes, NK cell eosinophils
Physical Barriers
Soluble Factors
Cells
none
Immunoglobulins(antibody)
T and B lymphocytes
ComponentsComponents of Innate and Adaptive of Innate and Adaptive ImmunityImmunity
PAMPs: Pathogen Associated Molecular Patterns
PRRs: Pattern Recognition Receptors
Takeda and Akira Genes to Cells (2001) 6:733-742
The NF-kB Family of Transcription FactorsThe NF-kB Family of Transcription Factors
Eukaryotic transcription factor found in essentially all cell typesEukaryotic transcription factor found in essentially all cell types First described in 1986 as a nuclear factor required for the First described in 1986 as a nuclear factor required for the
transcription of the immunoglobulin kappa light chain in B transcription of the immunoglobulin kappa light chain in B cells.cells.
Binds to a 10-bp sequence GGGGYNNCCYBinds to a 10-bp sequence GGGGYNNCCY Important component in the inducible expression of many Important component in the inducible expression of many
proteins: cytokines, acute phase proteins, adhesion moleculesproteins: cytokines, acute phase proteins, adhesion molecules The NF-kB signaling system is evolutionarily conservedThe NF-kB signaling system is evolutionarily conserved
Three NF-kB molecules in Three NF-kB molecules in DrosophilaDrosophila dorsaldorsal
controls dorsal/ventral polarity during developmentcontrols dorsal/ventral polarity during development Regulates antifungal gene expressionRegulates antifungal gene expression
difdif andand relishrelish: : regulate expression of antifungal and antibacterial regulate expression of antifungal and antibacterial genesgenes
NF-NF-B exists in the cytoplasm as an inactive B exists in the cytoplasm as an inactive heterotrimer composed of 2 Rel family proteins and heterotrimer composed of 2 Rel family proteins and
an inhibitory IkB moleculean inhibitory IkB molecule
IkB
p65 p50
IKK
Stress, infection, or cytokine
P P (Ub)n
26S proteosome
NuclearTranslocation
Activation of NF-KB Responsive genes
NF-NF-B Family StructureB Family Structure
Toll
Pelle
tube??
cactuskinase
cactus
dorsaldifrelish
Conservation ofConservation ofsignaling signaling pathwayspathways
between flies between flies and humansand humans
Mutation of genes in the Toll signalingcascade in Drosophila block the expression of anti-fungal and anti-bacterial genes
Water
A. fumigatus
E. coli
Signaling to monocytic cells through the human Toll homolog induces accessory cell functions
NF-B Activation
Requirements for the recognition of targets by the innate immune
Molecular structures recognized by the immune Molecular structures recognized by the immune system must be shared by large groups of pathogenssystem must be shared by large groups of pathogens molecular patterns vs. particular structures (antigens) PAMPmolecular patterns vs. particular structures (antigens) PAMP
PAMPs must be conserved products of microbial PAMPs must be conserved products of microbial metabolism not subject to antigenic variability metabolism not subject to antigenic variability
The recognized structures must be absolutely distinct The recognized structures must be absolutely distinct from self antigens: discrimination of self vs. non-selffrom self antigens: discrimination of self vs. non-self
LPS Binding Protein (LBP)LPS Binding Protein (LBP) 60-kDa serum glycoprotein that binds with high affinity to LPS Sequence homology to bactericidal/permeability increasing
protein Acute phase protein secreted by hepatocytes
Serum levels between 1 and 10 ug/ml in normal human serum
Concentrations >300 ug/ml in acute phase serum Critical for rapid responses to small amounts of LPS or gram-
negative bacteria and for survival of Salmonella infection Expression of LBP in hepatocytes is regulated by LPS, IL-1, IL-6
and TNF
Molecules involved in the recognition Molecules involved in the recognition of LPS by Cellsof LPS by Cells
Molecules involved in the recognition Molecules involved in the recognition of LPS by Cellsof LPS by Cells
CD14, sCD14CD14, sCD14 55 kDa GPI-linked protein on the surface on monocytes and 55 kDa GPI-linked protein on the surface on monocytes and
PMNPMN Also found as a soluble protein in serum:sCD14Also found as a soluble protein in serum:sCD14 Each CD14 molecule binds 1 molecule of LPS complexed to Each CD14 molecule binds 1 molecule of LPS complexed to
LBPLBP Required for responses of genes to low concentrations of LPS Required for responses of genes to low concentrations of LPS No cellular signaling activityNo cellular signaling activity
CD11b/CD18 (Mac1)CD11b/CD18 (Mac1) Alternative cell surface receptor for LPSAlternative cell surface receptor for LPS Recognizes LPS at high concentrationsRecognizes LPS at high concentrations
For expression of a fullFor expression of a full repertoire of LPS-inducible genes, CD14 repertoire of LPS-inducible genes, CD14 and CD11b/CD18and CD11b/CD18 must be coordinately engaged to deliver must be coordinately engaged to deliver optimal signaling tooptimal signaling to the macrophagethe macrophage..
Search for the LPS Gene Search for the LPS Gene Endotoxin hyporesponsive mice
1. C3H/He 1947 C3H/HeN: Normal LPS response
C3H/HeJ: LPS hyporesponsiveSpontaneous mutationmid 1960’s
2. C57BL/10SnSometimeAfter 1953
C57BL10/ScCRLPS hyporesponsive
January 1998 DNAX January 1998 DNAX reports the cloning of reports the cloning of human TLR1-5human TLR1-5 No ligands No ligands
describeddescribed Janeway’s Janeway’s
hToll=TLR4hToll=TLR4
Rock, et al. 1998 PNAS 95:588
September 1998 Genentech identifies TLR2 as the LPS signaling molecule by transfection studies Didn’t look at TLR4
December 1998 Tularik also identifies TLR2 as LPS signaling molecule in transfection studies TLR4 didn’t work
September 1998: Bruce Buetler’s lab mapped LPS to a 1.2Mb region of chromosome 4. The only intact gene within this region is TLR4
December 1998: “Defective LPS signaling in C3H/HeJ and C57BL/10ScCr mice: Mutations in Tlr4 gene” Poltarak et al. Science 282:2085
Poltarak et al 1998 Science 282:2085Poltarak et al 1998 Science 282:2085 A single nucleotide change in the gene for
TLR4 renders the C3H/HeJ mouse non-responsive to LPS
C3HHeN RFHLCLHYRDFI P GCAIAANIIQEGFHKC3HHeJ RFHLCLHYRDFI H GCAIAANIIQEGFHK
Hoshino et al 1999 J. Immunol. 162:3749Hoshino et al 1999 J. Immunol. 162:3749 TLR4 knockout mice are hyporesponsive to LPSTLR4 knockout mice are hyporesponsive to LPS
Dunne and O’Neil 2003 www.stke.org/cgi/content/full/sigtrans;2003/171/re3
ReceptorReceptor(Pattern Recognition (Pattern Recognition
Receptors)Receptors)
Agonist(s)Agonist(s)(Pathogen-Associated (Pathogen-Associated Molecular Patterns)Molecular Patterns)
TLR1TLR1 Heterodimerizes with TLR2Heterodimerizes with TLR2
TLR2TLR2 PGN, some LPS, some LTA, PGN, some LPS, some LTA, lipoproteins, AraLAMlipoproteins, AraLAM
TLR3TLR3 dsRNAdsRNA
TLR4TLR4 Gram(-) LPS, Taxol, some Gram(-) LPS, Taxol, some LTALTA
TLR5TLR5 FlagellinFlagellin
TLR6TLR6 Heterodimerizes with TLR2Heterodimerizes with TLR2
TLR7TLR7 ImidazoquinolineImidazoquinoline
TLR9TLR9 Bacterial DNA (CpG)Bacterial DNA (CpG)
TLR 8,10TLR 8,10 UnknownUnknown
Akira J.Biol.Chem. 2003 278:38105–38108
How do we explain the Genentech and Tularik How do we explain the Genentech and Tularik findings?findings?
How could TLR4 knockout mice be LPS-nonresponsive but How could TLR4 knockout mice be LPS-nonresponsive but TLR2 transfected cells LPS responsive? TLR2 transfected cells LPS responsive?
Why were TLR4-transfected cells LPS non-reponsive?Why were TLR4-transfected cells LPS non-reponsive?
1.1. In both cases, the LPS In both cases, the LPS preparations used by the preparations used by the investigators were investigators were contaminated with contaminated with bacterial lipopeptides: bacterial lipopeptides: TLR2 agonists!TLR2 agonists!
2.2. In Tularik’s experiments In Tularik’s experiments with transfected TLR4, with transfected TLR4, they lacked a critical they lacked a critical accessory protein required accessory protein required for efficient TLR4 for efficient TLR4 expression and function: expression and function: MD-2MD-2
0.0
1.0
2.0
3.0
4.0
5.0
6.0
7.0
8.0
9.0
TLR2Control
TLR2SigmaLPS
TLR2pureLPS
TLR2LTA
TLR2PGN
TLR4Control
TLR4SigmaLPS
TLR4pureLPS
TLR4LTA
TLR4PGN
Fo
ld In
du
ctio
n
0.0000
0.1000
0.2000
0.3000
0.4000
0.5000
0.6000
Control LPS PMA
Rel
ativ
e L
igh
t U
nit
s
Vector
MD2
MD-2
TLR4/4CD14IL-1RI IL-1RAcP
MyD88 MyD88
TIR domain
Death domain
IRAK IRAKTRAF6
TAK1/NIK
IKKComplex IkB
p65p50
Common Themes in IL-1, TLR, and IL-18 SignalingCommon Themes in IL-1, TLR, and IL-18 Signaling
IL-1RAcP
MyD88
IL-18R
IRAK
NF-B activation
Nat Immunol. 2002 Apr;3(4):392-8
Nature 2003 420:329
Nature 2003 420:324
TIRAP=MAL
www.stke.org/cgi/content/full/sigtrans;2003/171/re3Luke A.J. O’Neil
Science. 2003 Aug 1;301(5633):640-3
Barton and Medzhitov Science. 2003 Jun 6;300(5625):1524-5
MD-2
TLR4/4CD14TLR1/6TLR2
MyD88MyD88
IRAK IRAKTRAF6
TAK1/NIK
IKKComplex
IkBp65p50
Common and Distinct Themes in TLR SignalingCommon and Distinct Themes in TLR Signaling
TIRAP
TRIF
IFN-
TIRAP IRF3
MAP kinases
Rac
PI3K
AKT
TIR domain
Death domain
PI3K
CommonResponses
TLR4-Specific
TLR2-Specific
Single ligand-Single response vs. Multiple Ligands-complex response
Ozinsky and Underhill Current Opinion in Immunology 2002, 14:103–110
Other PRRsOther PRRs
Dectin-1Dectin-1 C-type lectin that binds b-glucan-containing particles including C-type lectin that binds b-glucan-containing particles including
zymosan and zymosan and C. albicansC. albicans Macrophage Mannose ReceptorMacrophage Mannose Receptor
Cell bound C type lectin that binds sugar molecules on the Cell bound C type lectin that binds sugar molecules on the surface on many bacteria and viruses.surface on many bacteria and viruses.
Macrophage Scavenger Receptor (SR-A)Macrophage Scavenger Receptor (SR-A) Recognize certain anionic polymers and low-density Recognize certain anionic polymers and low-density
lipoproteinslipoproteins Peptidoglycan Recognition Proteins (PGRP)Peptidoglycan Recognition Proteins (PGRP)
4 forms in humans:4 forms in humans: PGRP-L: liverPGRP-L: liver PGRP-Ia and PGRP-Ib: esophagus PGRP-Ia and PGRP-Ib: esophagus PGRP-S: bone marrowPGRP-S: bone marrow
AdjuvantsAdjuvants
Functionally defined as any substance that, when Functionally defined as any substance that, when mixed with antigen, increases the mixed with antigen, increases the immunogenicity of the antigenimmunogenicity of the antigen
Injection of unmodified, nonself proteins in the Injection of unmodified, nonself proteins in the absence of adjuvant results in tolerance.absence of adjuvant results in tolerance.