Transcript NOD-Like Receptors

Host-Pathogen Interaction
Sites of microbial infection
Mouth
Conjunctiva of eye
Skin
Pores, hair
Follicles,
Sweat glands
Scratch, injury
Lung
Insect bite
Digestive
tract
Urogenital
tract
Pathogenic microbes
Anus
Normal non-pathogenic
microflora
Pathogens can be successful
in causing an infection
 They can attach to and penetrate body surfaces
Ex: Schistosoma mansoni
 Pathogens can be introduced by a biting insect
Ex: Malaria, Leishmania, virus, bacteria
 Pathogens can take advantage of preliminary damage
(wound, respiratory tract damage)
Ex: Trypanosoma cruzi
Vector: Reduviid bugs (Triatoma & Rhodnius)
a.k.a. Kissing bug
However, the Immune System
has Natural, Constitutive Microbial Sensors
Utilized to Rapidly React to Invasive Pathogens.
They are part of the Innate Immune Response and
Play Pivotal Role in the Development of the
Adaptive Immune Response.
TLR Roles
O’Neill, Luke A.J. “Immunity’s Early-Warning System”. Scientific American, Jan (2005), 38-45.
O’Neill, Luke A.J. “Immunity’s Early-Warning System”. Scientific American, Jan (2005), 38-45.
Activation of adaptive immunity by
innate immunity
PGN
Lipopeptides
dsRNA
Unmethylated
Flagellin CpG DNA
LPS
TLR2
TLR4
TLR3
Cytokines (IL-1, IL-6, IL-12, TNF)
Chemokines (MIP-2, MIP-1a/b)
TLR5
TLR9
TLR2
TLR1
Co-stimulatory
molecules
CD28
B.7
NF-kB
Phagocytic
receptor
MHC
Phagosome
Microorganisms
TCR
Peptide
Naive T
Cell
Antigen Presenting Cell
Hemozoin: A Malarial Metabolic Waste
PfHZ
sPLHZ
Rapid crystalline (0.7-0.9 mm)
Size and Shape similar to PfHZ
(see Inset). Bar size is 200 nm.
Intracellular Microbial Sensors
Toll-like receptors:
Membrane Receptor that Sense Extracellular Microbes
and within phagosome/endosome.
NOD-Like Receptors, RIG and MDA:
Intracellular Microbial Sensors
Are Cytoplasmic surveillance proteins with CARD domain
NOD proteins (Nucleotide-binding oligomerisation domain)
Two members: NOD-1 and NOD-2
RIG-1 (Retinoid-induced gene 1)
MDA (Melanoma differentiation-associated gene)
CARD domain
CARD (Caspase-Activating and Recruitment Domain)
Found in some caspase proteins
Mediates protein-protein interaction
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RIG-I
1
MDA-5/Helicard 1
IPS-1/Cardif/VISA/MAVS
1
CARD CARD
Helicase domain
CARDCARD
Helicase domain
CARD
540
925
1025
Bouchier-Hayes L and Martin S.J., EMBO (2002)
NOD protein structure
CARD
NBD
LRRs
N-term
CARD
CARD
NBD
N-term
CARD (Caspase-activating and recruitment domain)
NBD (Nucleotide binding Domain)
LRRs (Leucine-Rich Repeats)
C-term
NOD-1
C-term
NOD-2
LRRs
NOD ligands: Peptidoglycan
Peptidoglycan (PGN)
Major component of gram+ cell wall
Found in thin layer in periplasmic space of gram–
Glycan chains alternating GlcNAc and MurNAc
linked by peptide bridge
MDP
Meso-DAP
Philpott D.J. and Girardin S.E., Mol Imm (2004)
NOD signalling
NOD2
PGN
NOD2
NOD1
RIP2
NOD1
RIP2
IKK
complex
RIP2
IkB
IkB
NFkB
Proteasome
NFkB
Transcription
NOD-Like Receptors (NLRs)
NOD1/2
IPAF/NAIP
NALP3/ASC
Bacteria
NOD-1
NOD-2
PGN
DAP (meso-diaminopimelic)
MDP (muramyl dipeptide)
PGN
Salmonella
Legionella
Flagellin
MDP
Francisella
RNA (PAMP)
Toxins
Danger-Associated Host Components
Uric Acid Crystals
Hemozoin ???
Meylan et al. Nature July 2006
(metabolic waste)
NOD-2 and Crohn’s disease
Abnormal NOD-2 expression correlates
with defective epithelial defense
 NOD-2 expressed in Paneth cells of intestine
 Enteric bacteria induce a-defensin through NOD-2
to kill luminal microbes.
 Clinical evidence: CD patients have decreased
a-defensin expression in Paneth cells
Summary
NOD-Like Receptors, RIG and MDA
Intracellular Microbial Sensors
That can detect bacterial and viral ligands
NOD detects PGN of several bacteria
Inflammatory response & Cell Death (IPAF/NAIP)
RIG-1/MDA-5 detects dsRNA of virus
Modulation of IFN signalling to control virus
Anti-microbial Functions
of Phagocytic Cells
Upon Receptor-Mediated recognition
Phagocytosis and Internalization
Phagosome Biogenesis
Anti-Microbial Products
Pro-Inflammatory Response (last lecture)
Major Players
Polymorphonuclear leukocytes (PMNs) / neutrophils
 Abundant, short lived
 Elevated number indicate infection
Professional antigen presenting cells
 Dendritic cells / Langerhan cells (skin)
 Monocytes (blood)  macrophages (motile or
stationary), which can be tissue specific
 Longer lived, lower frequency
Underhill and Ozinsky. Annu. Rev. Immunol. 2002
Formation of the phagolysosome
upon ingestion of a microbe
Phagosome maturation
Cellular Microbiology, 1999, 1(3):195-203
Degradation in the phagolysosome
Phagolysosome
Lysosome
Acid
hydrolases
Endosome
Bacteria
degradation
Phagocytosis and Anti-Microbial Products
Bacteria
Lysosomes
Phagolysosome
Plasma membrane
Phagosome
NAPDH
oxidase
O2 Proteases
Nitric Oxide
H 2 O2
Ionic strength
pH
TNF-a
+ TNF-R1
Induction of
iNOS
Expression
IFN-a/b + JAK/TYK
IL-1 + IL-1R1
LPS+TLR 4
IFN-g + JAK1/2
IKB
STAT1
STAT2
NFΚB
NFIRF-9
ΚB
ISREΚB
MAPKs
IL-6 + JAK1/2
SAPs/JNKs
p38
AP-1IRF-1
NF-IL6
AP-1GAS
NF-IL6
Citruline
Arginine
Active
Resting
O2
p22phox
O2-
Heme
gp91phox
FAD
NADPH
Rac
PKC
PI3K
p40phox
P
P p47phox
PKA
P
P
MAPK
P67phox
Reactions Between ROS and RNS
NO + O2-
NO2 + ONOO-
NONOates
S-nitrothiols
Nitrite
Nitrous Acid
Chakravortty and Hensel. Microbes Infect. 2003.
Mechanisms of Microbicidal Activity
Polyunsaturated Lipids
DNA
Formation of Oxylipins disrupts membrane
Enzyme Deamination
Function of nucleosides
Abasic sitesand Nitrosylation of SH groups
Oxidization
Strand breaksof Tyrosine residues
Nitrosylation
Inactivation of metal ions at active site
Depletion of antioxidants
Evasion:
“the act of physically escaping from something (an opponent or a
pursuer or an unpleasant situation) by some adroit maneuver”
Nature 3:11, 2002 editorial
•Passive
•Active
Bacterial defense against phagocytes
Green: host
Orange: bacterial
 √ Induction of apoptosis
 X Uptake
√ Down-regulate 
√ Escape 
X Endosomal
trafficking/
Phagosome
maturation 
X Defense
factors 
EVASION MECHANISMS
Streptococcus suis type 2
Wild type S735
Mutant 2A
Phagocytosis
resistance
Phagocytosis
sensitive
Adapted from Charland et al.,1998
MS150502
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Host Signalling Alteration
Acid
phosphatase
Ca2+
Channel
Ca2+Influx
LPG
[Ca2+]i
PKC
PMA
IFNg
Tyrosine
phosphatase
(SHP-1)
JAK1
PTK
Ser⁄Thr phosphatase
JAK2
DAG
Protein phosphorylations
PIP2
FMLP
R
G
IP2
STAT
Ca2+
PLC
INT
EXT
Ca2+stores
Cellular functions
• H2O2,O2• IL-1
• phagocytosis
• MHC Class II
• c-fos expression
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