These slides provide information about mechanism of innate immunity and receptors recognizing pathogens such as TLRs, NLRs, CLRs etc.

1. Innate Immunity and Adjuvants
Sukhbir Kaur

2. History of Immunology
• Discovery of vaccination
by Edward Jenner in
1796.
• Based on observation
cowpox infections
protected the body
against small pox.
• Discovery of antibodies
and passive immunity
by Shibasaburo
Kitasato and Emil von
Behring in 19th century.

3. • Paul Ehrlich - side chain theory
of immunity.
• It predicted mechanism of
antibody production from B-
cells.
• Discovery of process of
phagocytosis by Elie
Metchnikoff (1895-1916).
• Ehrlich and Metchnikoff togather
got Nobel Prize for Medicine for
discovery of humoral(adaptive)
immunity and innate immunity
respectively.

4. Immune responses
Immune system
Adaptive immunity
Cell-mediated
T-cells
Humoral
B-cells
Innate immunity
Macrophages,
Dendritic cells and
leucocytes.

5. Adaptive immunity
Main components: B-cells and
T- cells which carry receptors
known as BCR and TCR.
Receptors have variable region
and constant regions.
Different gene segments
encode variable regions.
Each member of gene segment
is randomly joined to other
members. This leads to huge
diversity of receptors.
Receptors reacting with host
components are eliminated.

6. Invasion by
pathogen
Activation of T-cells and B-cells
Activated cytotoxic
and helper T-cells
Antibody producing
Plasma cells
Memory T-
cells Memory B-cells.

7. Innate immunity
• Main components : Leucocytes ,Macrophages
and Dendritic cells (collectively known as
phagocytes).
• Not long lasting.
• Immediate and temporary system of defence.
• Discriminate self and foreign components. This
ability relies to greater extent on TLRs .
• Activation of innate immunity is necessary for
induction of adaptive immunity.

8. Innate and adaptive immunity

9. Two theories of immune responses
Original theory
• Dendritic cells engulf
pathogen, digest it into
peptides and express these
peptides on their surface.
• Migrate from site of infection
to lymph nodes and present
antigens to naïve T-cells with
corresponding receptors .
• Pathogen is recognized in
lymph nodes during T-cell
activation.
New theory
• Activation of dendritic cells
is needed to activate T-cells.
• Dendritic cells remove dead
cells from host and self
antigens are presented to T-
cells but no activation of T
cells as dendritic cells are
not activated.
• Activation is mediated by
TLRs.

10. Toll like receptors (TLRs)
• Evolutionarily conserved
between insects and vertebrates.
• Type I glycoprotein having :
• extracellular domain
characterized by varying no. of
leucine rich repeats(LRR) motifs.
• Cytoplasmic signaling domain,
homologous to that of
interleukin(IL)-1 receptors,
known as Toll/IR-1 (TIR) domain.
• LRR domains consist of 19-25
tandem repeats of 24-29 amino
acids and these are xLxxLxLxx
acid residues.
• LRRs have horse shoe structure
and are involved in ligand
recognition

11. TLRs
• The ligands for
TLRs:
a. lipids,
b. protein and
c. nucleic acids
• These are potent
immune adjuvants
that can trigger
vigorous immune
response so TLRs
are called adjuvant
receptors.

12. TLR Ligand Notes
TLR1+TLR2 Bacterial lipoprotein(BLP)
(triacylated)
Immune and non-
immune cells
TLR2+TLR6 Mycoplasma macrophage
activating lipopeptide-2
(MALP) (diacylated),
Immune and non-
immune cells.
TLR3 Viral DNA and RNA
TLR4 Lipopolysacchrides(Gram
–ve bacteria)
Most potent ligand
TLR5 flagellin Digestive tract,
respiratoory tract, urinary
tract, dendritic cells in
mucosa
TLR7, TLR8 Imidazoquinolin,
nucleoside guanosine or
uridine rich ssRNA
TLR8 not active in mice
TLR9 Unmetylated CpG motifs Bacteria have
unmethylated CpG

13. TLR Ligands and TLR4

14. TLR signaling pathways
• Adaptor molecules which interacts with TIR
domain of TLRs to activate downstream signaling
pathways:
• MyD88: in all TLRs except TLR3. So, 2
pathways:MyD88 dependent and independent.
• TRIF :used by TLR3 and TLR4.
• TIRAP: used by TLR2 and TLR4 to recruit MyD88.
• TRAM: bridge between TLR4 and TRIF.
MyD88 NF-κB and MAP kinase activation
inflammatory cytokines.
TRIF NF-κB andIRF3(interferon regulatory
factor 3) activation type I IFN.

15. TRIF dependant
signaling involves
TANK-binding kinase
1(TBK 1) and IκB
kinase, inducible
(IKKi) acting asIRF3
kinase

16. RLR- cytoplasmic helicases
• Retinoic acid-inducible gene I(RIG-I)- RNA
helicase, present in cytosol, detects viral
infections.
• Melanoma differentiation-associated gene 5
(MDA5) is homologous to RIG-I.
• Both are found ubiquitously in all cells.
• Possess two N-terminal caspase-recruitment
domains (CARDs) followed by helicase domain.
• RIG-I 5’-triphosphate short or long
dsRNAs.
• MDA5 picornavirus family(has long
dsRNA of more than 2kbp.

17. RLR: signaling pathway
• CARDs responsible
signal transduction
which leads to NF-κB
and IRF3/7 activation
via adaptor molecule
IPS-1 (IFN-β promoter
stimulator 1)
• For IRF3/7 activation ,
TBK1 and IKKi are
involved.
• For induction of type I
IFN, STING (stimulator
of interferon genes)
also acts as a mediator.
It is multi spanning
membrane protein that
associates with TBK.

18. NLRs
• NLRs consist of: C-
terminal LRR domain,
central NOD domain and
N terminal effector
domain which is involved
in signaling.
• NOD1 and NOD2 –
recognise components of
peptidoglycan activate NF-
κB and induce
inflammatory response
• Other NLRs are involved in
inflammosome formation
and production of mature
IL-1β and IL18 .

19. For inflammosome formation ,NODs activate Caspase-I
which cleaves pro IL-1β to mature IL-1β .
4 types of inflammosomes:
 nucleotide-binding domain and leucine-rich repeat
containing family pyrin domain containing1 (NLRP1)
 nucleotide-binding domain and leucine-rich repeat
containing family ,CARDdomain containing4 (NLRC4),
 NLRP3 and
 absent in melanoma 2 (AIM2).
 NLRC4 detects flagellin and imidazoquinoline
 NLRP3 detect monosodium urate, abestos and
cholesterol crystals.
NLRs

20. CLRs
 C- type lectin receptor (CLR) recognize
β- Glucan which is cell wall
component of fungi and yeast.
 It consists of :
 Extracellular carbohydrate
recognising C- type lectin domain-
CTLD
 cytoplasmic tyrosine containing
domain
 Dectin-1 recognizes Candida,
Aspergillus, Pneumocystis and
Coccidioides .
 Interaction of Dectin-1 with β-Glucan
triggers spleen tyrosine
kinase(Syk)which activates NF- βB
through CARD9,Bc110 and MALT1 as
well as caspase-1.

21. CLRs
• Dectin -2 expressed on: tisue macrophages,
dendritic cells and inflammatory monocytes.
• It has a classical sugar binding CTLD which
recognizes high mannose structures in Ca2+
dependent manner.
• It recognizes: Cryptococcus neoformans, Candida
albicans, Saccharomyces cerevisiae,
Mycobacterium tuberculosis,
Microsporumaudounii, Paracoccidioides
brasiliensis and Histoplasma capsulatum.

22. Induction of adaptive immunity

23. Conclusion
• For development and modulation of innate
immunity, innate immunity is necessary.
• PRRs-TLRs, RLRs, NLRs and CLRs recognize
pathogens and inflammatory reactions which
trigger adaptive immunity