Overview of Innate Immunity

Question 1

Define the innate immune system

Answer 1

Immune response responsible for the confinement of infectino in the early hours of exposure.

Depends on the recognition of molecular patterns found in micro-organisms

Question 2

Compare the innate and adaptive immune system with regards to:

Time

Stimulus

Memory cells

Diversity

Organism

Answer 2

Question 3

Name 7 cells involved in the innate immunity of the lungs (name some molecules associated with them and/or describe their main function)

Answer 3

1. Dendtritic cells - TLRs, IFNs, cytokines
2. Neutrophils - phagocytosis (ROS)
3. Innate Lymphoid Cells (ILCs)
4. NK and NKT cells
5. Macrophages - phagocytosis and scavenging
6. Airway epithelium - IL-25, IL-33, TSLP
7. γδ T cells - TLRs

Question 4

Briefly describe the function og dendritic cells

Answer 4

Sample the airway mucosa and releases cytokines, chemokines, interferons as a response. It links the innate and adapive signalling together

Question 5

Where are Innate Lymphoid Cells (ILCs) found and how are they different to T-cells

Answer 5

Sentinel cels found at mucosal surfaces.

Same effector functions to T-cells but lack TCR (T-cell receptor

Question 6

Describe the three groups of ILCs, their function and cytokines they produce

Answer 6

ILC1 = Th1 (IFNγ, TNF-α); intracellular microbes

ILC2 = Th2 (IL-4, IL-5, IL-9, IL-13); allergic big cytokine producer

ILC3 = Th17 (IL-17A, IL-22); extracellular microbes

IL-25, IL-33, TSLP (epithelial cytokines) stimulate these ILCs

Question 7

Epithelial cytokines are considered _____ because they are preformed - they are ready to be released in response to an insult. Three alarmins are: ______. Epithelial cells are mechanically sensitive

Answer 7

Epithelial cytokines are considered alarmins because they are preformed - they are ready to be released in response to an insult. Three alarmins are: IL-25, IL-33, TSLP. Epithelial cells are mechanically sensitive​

Question 8

Outline the kinetics of the immune response to acute viral infections, with regards to viral load, IFN, NK cells, T cells, IgG

Answer 8

Viral load increases from exposure (peaks day 2 and drops day 5)

There is a lag phase between the viral load increase and the IFN increase (INNATE)

NK cells increase follows IFN

Adaptive immune response is late (~day 4) (T cell increase)

IgG and IgA are produced much later (sharp increase at day 11)

Question 9

Compare the immune response to RSV in mice, adults, infants and elderly.

Answer 9

Response to pathogens different and changes rapidly with age and co-morbidities

Question 10

Briefly outline the role of defensins and cathelicidins

Answer 10

Broad bactericidal and anticiral properties - non-specific perforation of pathogen membranes (similar to complement system)

Question 11

Lactoferrin (transferring family) secreted by _____ and _____ cells. It’s main function is the prevention of bacteria proliferation by _____ of _____ which bacteria rely on for oxygen capture (has best influence in anaerobic conditions).

It also has other functions:

1. _____: binds to surface glycoproteins (HIV, CMV)
2. Bactericidal: _____ formation on Candida sp.
3. Bacteriotoxic: prevents _____ formation

Answer 11

Lactoferrin (transferring family) secreted by neutrophils and epithelial cells. It’s main function is the prevention of bacteria proliferation by sequestration of oxygen which bacteria rely on for oxygen capture (has best influence in anaerobic conditions).

It also has other functions:

1. Anti-viral: binds to surface glycoproteins (HIV, CMV)
2. Bactericidal: Pore formation on Candida sp.
3. Bacteriotoxic: prevents biofilm formation

Question 12

Name and describe the function of 4 Pattern Recognition Receptors (PRRs)

Answer 12

1. Toll-like receptors (TLRs) - recognise PAMPs
2. NLR - recognise dsRNA, muramyl dipeptides and peptidoglycans
3. RNA helicases (RIG-I) - recognise intracellular viral dsRNA and ssRNA
4. C-type lectins - recognise carbohydrates

NLR = nucleotide-binding oligomerization domain (NOD) like receptors

Question 13

Name two molecules recognised by PRRs and describe their features.

Answer 13

1. PAMPs (Pathogens) - non-self, DNA form, dsRNA, bacterial cell wall products
   
   1. invariant molecules (products that cannot be mutated to avoid recognition)
2. DAMPs (damage) - self - normal molecules but are found in laces they should not be (ATP). Can be modified-self DAMPs (citrullinated peptides act as auto-antigens and generated during inflammation)

Much of the signalling appartus is shared between PRRs and IL receptros (IRAKs)

Question 14

Nam 4 TLRs found in cellular endosomes and name their ligand(s)

Answer 14

1. TLR3 - dsRNA
2. TLR7 and TLR8 - ssRNA
3. TLR9 - unmethylated CpG DNA

TLRs in recognising viral invariants

Question 15

Name two TLRs found exclusively in the cell surface membrane

Answer 15

TLR4

TLR5

Question 16

What do NOD-like receptors (NLRs) recognise

Answer 16

dsRNA

peptidoglycans

muramyl dipeptides

(i.e. bacterial cell wall products)

Question 17

Where are RNA helicasesor RIG-like helicases(RLHs) found?

Answer 17

In the cytosol/cytoplasm

Question 18

RIG-like helicases are characterised by what?

Answer 18

DEAD box protein - conserved motif: Asp-Glu-Ala-Asp (DEAD)

Question 19

Name three RIG-like helicases, describe their structure and name their ligands/function

Answer 19

1. RIG-I - has RNA helicase domain and CARD. short dsRNA and 5’phospho ssRNA
2. MDA5/Helicard - has RNA helicase domain and CARD. Lond ds RNA
3. LGP2 - RNA helicase domain. Inhibitory helicase (no CARD)

RIG-I = retinoic acid inducible geneI; CARD = Caspase Recruitment Domain; MDA5 Melanoma differentiation associated gene; LGP2 = Laboratory of genetics and physiology 2

Question 20

Outline in 3 steps the PRR signalling

Answer 20

1. Receptor
2. Adaptor
3. Transcription Factor

Question 21

What is needed when RIG-I and MDA5 bind to their ligand to go through conformational shpe change

Answer 21

ATP

Question 22

Name the adpator protein that RNA helicase uses for signalling

Answer 22

VISA/CARDIF/MAVS/IPS

Same adaptor, four different names

_V_eronica _C_an’t _M_ove _I_n

Question 23

What Transcription factors do RIG-I and MDA5 (RNA helicases) activate and what proteins o they upregulate

Answer 23

NF-κB - pro-inflammatory (TNF, IL-8)

IRF-3 (1, 3, 7)- interferon, antivirals (IFN-B)

IRF-3: Interferon Regulatory Transcription Factor 3

Question 24

Name the signalling complex associated with activation of IRF-3 TF by the RIG-5/MDA5 pathways

Answer 24

TANK

IKK-ι/ε

TBK-1

TBK = TANK binding kinase

Question 25

Name the signalling complex associated with activation of NF-kB TF by the RIG-5/MDA5 pathways

Answer 25

NEMO IKK-α IKK-β

Question 26

Describe the complete PRR signalling involved in IFN-β transcription

Answer 26

1. Viral material enters cell (TLR4) 2. Material in **cytoplasm** (recognised by RIG-I and MDA5) or in **endosome** (TLR3, 7, 8, 9) 3. Adaptor protein binding (VISA/CARDIF/MAVS/IPS) 4. Signalling complex binds (TANK/IKK-_i/e/TBK-1) 5. Phosphorylation and activation of IRF-3 and binding to nuclear binding site to upregulate the **transcription of IFN-β** (NOT expression)

Question 27

Decribe the downstream effects of secreted IFN-β in neighbouring cells

Answer 27

1. IFN-β binds to its receptors (IFNAR1/2) 2. Activation of kinase cascade involving JAK1/2 and STAT1/2. 3. Phosphorylation of IRF7 and IRF9 and binding to nuclear binding site 4. Increase transcription of **Interferon Stimulated Genes** (ISGs - e.g. OAS and RNAse L) ## Footnote *Interferons are **not antivira** - it is their effects on target cells which mediates antiviral response*

Question 28

How many types of human interferons are there and provide examples with what type of cells secreting these IFNs

Answer 28

1. **Type I**: IFN-β (all nucleated cells) 2. **Type II**: IFN-γ (**strictly** leukocytes) 3. **Type III**: IGN-λ (most nucleated cells)

Question 29

Name 5 Interferon stimulated genes and theri function

Answer 29

1. **PKR**: binds viral **dsRNA** and stops host cell **translation** 2. **Viperin**: prevents viral protein **trafficking** 3. **MxA**: GTPase preventing viral **replication** 4. **RNAseL**: degrades viral dsRNA 5. **OAS**: activates RNAse L with 2'-5' oligoadenylate *OAS - 2'-5' oligoadenylate synthetase*

Question 30

Describe the mechanism of function of RNAse L and OAS ## Footnote *OAS = 2'-5' oligoadenylate synthetase*

Answer 30

1. IFN-B signalling upregulates the transcription of IFN stimulated genes (ISG) 2. OAS is one of these ISGs and it recognises viral **dsRNA**. 3. OAS activates and arms RNAse L with 2-5A. 4. Active RNAse L degrades the RNA, thereby blocking replication

Question 31

Describe 6 effects of IFN-β on the immune system

Answer 31

1. Induce **anti-viral state** through **ISGs**(OAS and RNAse L) 2. ↑other IFNs (IFN-αs) 3. ↑RNA helicases (↑IFN-β transcription) 4. T Cell cytokines (CCL5, CXCL10) 5. Apoptosis of infected cell 6. Activates **dendritic cells** ## Footnote *These are energetically expensive processes and damage host cell replication - so they are **inducible***

Question 32

Dendritic cells link the innate and adaptive immune response. Name the two types of dendritic cells and their function

Answer 32

1. **Plasmacytoid DCs** (pDCs) 1. Peripheral (circulation) 2. IFN-α/β producers 3. ↑PRRs (TLR7/8/9) 4. Poor activation of Th1 cells 2. **Conventional DCs** (cDCs) 1. In tissue 2. Activators of Th cells 3. ↑integrins and PRRs 4. *Further subdivided into CDC11c*

Question 33

Describe the three main ways in which dendritic cells link innate and adaptive immunity

Answer 33

1. PRRs recognise PAMPs 2. Respond to signals from infected cells 3. Mediate T cell function

Question 34

Explain how RNA interfaces (RNAi) can regulate/silence gene expression

Answer 34

siRNA or miRNA (RNAi) are delivered or naturally produced in the cell, respectively DICER cleaves siRNA and miRNA into smaller sections AGO binds to these and degrades one strand The ss and AGO complex is called RISC RISC finds its match (siRNA - perfect; miRNA seed matching therefore lots) Cleaves at point of match

Question 35

Briefly describe 4 virus/gene or protein which counterats the innate immune response

Answer 35

1. Modification of host proteins (Vaccinia B8R is IFNAr1 homolog - doesn't function) 2. Hep C cleaves CARDIF/IPS-1 3. Influenza/ NS1 complexes with RIG-I and prevents ligation with adaptors (CARDIF-IPS)

Question 36

Describe the features of a deficiency in IFN-β and IFN-γ

Answer 36

HIgher viral load greater decrease in lung function

Question 37

Describe the features of IL-1 Receptor Associated Kinase (IRAK)-4 - an adapter kinase downstream of TLRs

Answer 37

↓produce inflammatory cytokines ↓IFN-a/B and IFN-λ Recurrent infections with pyogenic bacteria normal immunity to viruses