Innate immune mechanisms Flashcards
How are pathogens detected?
T and B cells- Individual T and B cells are highly selective for a specific pathogen species.
Immune memory – faster immune response on 2nd infection by the same pathogen.
Innate immune cells- Less selective-recognizes broad classes of pathogens and not specific species.
No immune memory.
1st line of defense.
What are pattern recognition receptors and what do they do?
Innate immune cells detect pathogens via pathogen associates molecular patterns (PAMPs)- Molecules or structures that are specific for microbes and viruses and not visible on host cells.
PAMPs are recognized by “pattern recognition receptors” (PRR):
- Unlike T cell or B cell receptors, PRRs are invariant germline encoded receptors.
- Expressed on innate immune cells and in some cases T cells, B cells and endothelial cells.
Several classes of PRR exist:
Membrane receptors- Toll like receptors, C- type lectin receptors.
Cytoplasmic receptors- Nod like receptors, DNA receptors, RNA receptors (RIG-I, Mda5).
How does loss of TLR4 protect mice from endotoxic shock?
Systemic bacterial or fungal infections result in sepsis:
- Excessive production of pro-inflammatory cytokines.
- Massive inflammation leading to organ failure and death.
- Mortality driven by over activation of the immune system rather then the invading pathogen. Response to pathogen, not pathogen itself, that will kill you.
Injection of bacterial endotoxin can give rise to a similar condition where cytokine driven inflammation results in endotoixc shock and death.
LPS into a normal mouse will result in death.
LPS in a TLR4 mutant or knockout mouse will do nothing.
Can give knockout same amount of LPS as normal (which could be a lot), and nothing will happen.
How does the immune system interact with pathogens?
Direct pathogen recognition by innate immune cells.
Direct pathogen recognition by adaptive immune cells (T and B cells).
Cell/cell communication and cross activation between innate and adaptive cells.
Leads to inflammation and pathogen killing.
What are the functions of TLRs and what is their outcome?
Phagocytosis, production of rective oxygen species- leads to pathogen killing.
Production of prostaglandins, production of chemokines, production of pro-inflammatory cytokines- leads to recruitment of immune cells to the infection site, and activation of other immune cells.
How are TLRs used in disease?
Immune activation is critical to clear pathogens:
Infection – TLRs play important roles in activating the host response to pathogens.
IRAK4/Myd88 mutations give rise to increased susceptibility to bacterial infections.
- Occurs in childhood; 30-40% mortality even with antibiotic treatment.
- Severity decreases with age.
TLR3 / Traf3 mutations result in recurrent herpesviris encephalitis.
Cancer – TLR activation occurs during the killing of cancer cells by immune cells; this process is subverted by cancer cells during tumor development.
Chronic inflammation underlies pathology in a large number of diseases:
Aberrant activation of TLRs can contribute to the development of autoimmune disorders.
SLE frequently associates with an inability to clear dead cells.
- Release of DNA from dead cells can activate TLRs leading to inflammation.
- TLR activation helps drive the production of autoantibodies.
Rheumatoid Arthritis-
Increased activation of multiple TLRs.
Diabetes- Associated with chronic inflammation; TLR driven activation of macrophages present in the adipose tissue have been suggested to be responsible for this.
Cardiovascular disease- Increased TLR activation contributes to the development of foam cells – specialized macrophages that are present in atherosclerotic plaques.
(Main role- response to infection. This would be compromised if these were absent.
Adaptive IS starts to compensate in teens.
TLR response v important in early childhood.
Lipids can activate TLR activation- leads to inflammation.)
What are toll like receptors?
Part of the IL-1 receptor family.
The number of TLR genes is species dependent: 10 in human, 12 in mice.Purple sea urchin has 253 TLRs.
Different Toll like receptors are found if different locations- some are on the cell membrane- respond to different things, some are in the endosome- respond to nucleic acids.
Different TLRs recognize different types of PAMPs.
Different classes of pathogen express types of TLR agonists.
Endogenous ligands exist for several TLRs:
- “Danger associated molecular patterns” or DAMPs.
- Can give rise to unwanted inflammation.
Correct cellular localization of TLRs is important:
- Must be in a position to bind to see the pathogen.
- Must be in a localization that minimizes exposure to endogenous host ligands.
(TLR9 is a good example- recognise DNA. If in membrane, could recognise DNA from other host cells or DNA nets from immune cells- give an unwanted response.
CpG sequence often methylated in human cells, but not in bacteria.)
What is the structure of TLRs?
Two parts (dimer)- arranged into C shaped curves.
Extracellular and intracellular domain.
Extracellular is a leucine rich repeat domain for ligand binding. Binds in middle of curve- not always direct, so accessory protein sometimes binds to help present properly.
Intracellular is a TIR domain for activation of signalling. Also found in IL-1 receptor. Mediate interaction between extracellular part and signalling.
TIR: Toll/interleukin-1 receptor homology domain.
Found in both TLRs and IL-1 receptors.
Also present in TLR/IL-1 adaptor proteins.
Mediates the interaction between TLRs and adaptor proteins; critical to initiate downstream signaling.
Toll like receptor signailling?
TLRs activate signaling via either Myd88 or Trif dependent signaling. Adaptors proteins associate with TLRs.
All TLRs except TLR3 couple via Myd88- ForTLR2 and TLR4 Myd88 recruitment is promoted a 2nd adaptor, Mal.
TLR3 and TLR4 can couple via Trif- Recruitment of Trif to TLR4 is mediated by Tram.
Describe MyD88 dependent signalling.
Mal and 88 have TLR domains too.
Mal binds to TLR. Other end of 88 binds IRAK4. IRAK4 is recruited to Myd88 via interactions between the death domain of Myd88 and IRAK4.
Recruit 2 more kinases. IRAK4 recruits and phosphorylates IRAK1 and IRAK2; This activates the kinase activity of IRAK1. IRAK2 lacks key catalytic residues in the kinase active site and is inactive. Kinase dead mutants of IRAK1 can mediate signaling, suggestion the major role of IRAK1 and 2 is as scaffold proteins.
Death domains mediate recruitment.
4 phosphorylates 1, this activates signalling complex. 2 has an inactive kinase domain- primarily acting as scaffold.
All irak mediate recruitment of Traf6.
What is IRAK4?
IRAK4 has a death domain at N end and a kinase domain at C end.
Death domain- 6-alpha helical bundle, Mediates protein-protein interactions.
IRAK4 is essential for Myd88 dependent signaling.
Knockout of IRAK4 in mice blocks responses to LPS and increases infection.
Inactivating mutations in humans results in the development of invasive bacterial infection.
Can be treated by prophylactic antibiotics in childhood; infections decrease with age.
What do IRAK1 and 2 do?
IRAK1 and 2 promote the formation of a ubiquitin signaling complex.
IRAK1/2 interact with Traf6.
This results in the addition of Lys63 polyubiquitin chains to both Traf6 and IRAK1.
While Traf6 has E3 ligase activity, the physiological E3 ligase in TLR signaling in not clear.
Lys 63 chains are added by Ubc13/UVE1A.
Linear polyubiquitin are the added by the Lubac complex.
Formation of chains critical to downstream signalling.
Ub domains in TLR signaling?
The formation of Lys63 and linear polyubiquitin chains is necessary for the recruitment and activation of key signaling enzymes in TLR signaling.
Blocking the formation of these chains by via knockout of the required E2 ligases in mice prevents the activation of downstream signaling events following TLR activation.
Key ubiquitin binding domains:
- NFZ – binds Lys63 polyubiquitin chains.
- UBAN – binds linear polyubiquitin chains.
Chains important to recruit the downstream enzymes.
What happens in the activation of Tak1?
Tak1 complexes consist of the kinase Tak1 and the accessory subunits Tab1, Tab2 and Tab3.
Tak1 is in the MAPK Kinase Kinase family.
Tab1 pseudophosphatase and is involved in regulating Tak1 activity in the cell.
Tab2 and Tab3 have an C-terminal NFZ domains that bind specifically to Lys63 polyubiquitin.
Tak1 key component of this- kinase that mediates activation of pathways.
What makes up the NFkB pathway?
The NFkB pathway is made up of the NFkB transcription factor, IkB inhibitory proteins and K KappaB Kinases (IKK)s.
Activated downstream of multiple stimuli not just TLR signaling.
Multiple activation mechanisms including the “canonical” activation pathway that is involved in TLR signaling.
Activation of cytokines, but have other roles outside of the IS.
