Innate Immunity III

What does every single cell in our body have the ability to do?

Recognize a viral infection and make type I IFNs

Describe the steps for intracellular recognition of a viral nucleic acid

1. Virus gets into a cell

2. Viral replication takes place

3. Virus will have an uncapped RNA with a 5’ triphosphate (5’ triphosphate RNA)

4. This nucleic acid is not mammalian which leads to the helicase domain of RIG-1 (protein the cytoplasm) to bind to it

5. Once bound the 5’-triphosphate will go to the mitochondria

6. At the mitochondria it will bind to MAVS

7. MAVS will activate TRAF6

8. TRAF6 leads to the initiation of transcription factors IRF3 and IRF7

9. Once IRF3 and IRF7 are phosphorylated they are activated and will go to the nucleus

10. At the nucleus they will initiate the transcription of IFN-B and IFN-a

11. IFN-B and IFN-a will then be secreted from the cell

What are the two methods that IFN-a and IFN-B can be produced?

1. Autocrine

2. Paracrine

Describe the autocrine process in virus-infected cells

1. Virus infects a cell and replicates

2. Intracellular recognition of a viral nucleic acid takes place as described above

3. IRF3 will go to the nucleus and bind to the promoter of IFN-B

4. NFkB and AP-1 also contribute to the binding of the promoter

5. IFN-B is produced and secreted from the cell

6. IFN-B will bind to a Type I IFN receptor on the surface of the same infected cell

7. Eventually IRF7 is produced

8. IRF7 will go to the nucleus and bind to the promoter of IFN-a

9. IFN-a will then be produced and secreted from the cell

Describe the paracrine process in virus infected cells

1. IFN-B is secreted from the virus infected cell

2. IFN-B will bind to type I IFN receptor on an uninfected cell

3. IRF3 is produced and will bind to the promoter of IFN-B in the nucleus

4. IFN-B is then produced and secreted

What cells can make IFN-a?

Immune cells, virally infected cells, and structural cells

What transcription factor is associated with IFN-a? IFN-B?

IFN-a: IRF7

IFN-B: IRF3

What is autocrine? Paracrine? (in terms of Type I IFNs)

Autocrine is where the production of IFN-B in a virus infected cell will stimulate the production of IFN-a in the same cell. While paracrine is IFN-B from a virus-infected cell will stimulate the production of IFN-B in an uninfected cell.

What are the genes turned on by interferon signalling called?

Interferon Stimulated Genes (ISG)

Describe the structure of an IFN receptor

Is a heterodimer made of interferon alpha receptor 1 (IFNAR1) and interferon alpha receptor 2 (IFNAR2)

Describe the process for ISGs being turned on

1. IFN-a or IFN-B can bind to Type I IFN receptor

2. Once bound, TYK2 and JAK1 will bind to the receptor, which causes them to be phosphorylated

3. TYK2 and JAK1 will phosphorylate STAT1 and STAT2

4. STAT1 and STAT2 will dimerize

5. The dimerized STAT will migrate to the nucleus and will bind to the promotor of different ISGs to initiate the transcription

What are the four different ISGs?

1. Protein kinase R

2. 2’,5’-oligo(A) synthetase

3. Mx proteins

4. IFIT proteins

What are TYK2 and JAK1?

They are kinases (can phosphorylate themselves)

Where are STAT1 and STAT2 found? Are they found in an activated or inactivated state?

In the cytoplasm (inactivated state)

Describe what happens when protein kinase R is transcribed

1. Protein kinase R will bind to dsRNA

2. Elongation factor 2 (elF2a) will be phosphorylated

3. Leads to the inhibition of translation

What is a hallmark of viruses? Why?

dsRNA. This is because mammalians do not have this

Describe what happens when 2’,5’ oligo(A) synthetase is transcribed

1. 2’,5’-oligo(A) synthetase will bind to dsRNA

2. Oligo A is then made

3. Oligo A binds to RNase L

4. RNase L will chew up RNA (leads to mRNA degradation)

Describe what happens when Mx proteins are transcribed

1. Mx proteins are phosphorylated by the hydrolysis of GTP

2. Mx proteins will then polymerize

3. This results in the inhibition of virus transcription and assembly

Describe what happens when IFIT proteins are transcribed

1. IFIT proteins will bind to dsRNA

2. This allows the IFIT proteins to bind to elF3

3. Results in the inhibition of translation

What do the products of ISGs generally do?

1. Inhibit transcription or translation of all genes in the cell which ultimately prevents viral genes from being used

What cells can perform phagocytosis?

1. Neutrophils

2. Macrophages

3. Dendritic cells

What are two ways that phagocytosis can be initiated?

Binding to microbes via:

1. Bound antibodies (opsonization)

2. Scavenger receptors

What is phagocytosis enhanced by?

The pathogen binding to PRRs

What is a way that phagocytosed microbes are killed?

A respiratory burst will generate reactive oxygen and nitrogen intermediates

How can activated neutrophils and macrophages kill phagocytosed microbes?

1. Express inducible nitric oxide synthase that produces nitric oxide with potent antimicrobial activity

2. Use hydrolytic enzymes and antimicrobial peptides (non-oxidative)

Describe the process of phagocytosis

1. Bacterium binds to Fc receptors for antibodies or PRRs on membrane evaginations called pseudopodia

2. Bacterium is ingested forming a phagosome

3. Phagosome will fuse with a lysosome

4. Bacterium is killed and then digested by low pH-activated lysosomal enzymes

5. Digestion products are released from the cell

6. The peptides produced from the cleavage of microbial proteins are loaded onto MHC Class I or II proteins to help with T activation

What is SR-A1? Does it induce phagocytosis? Does it signal? What are the target pathogens? What are the ligands?

It is a scavenger receptor. Yes it initiates phagocytosis. It does not signal. Bacteria, Hep C virus are the target pathogens. Ligands are LPS, LTA, proteins, CpG DNA

What is SR-A6? Does it induce phagocytosis? Does it signal? What are the target pathogens? What are the ligands?

It is a scavenger receptor and it does initiate phagocytosis. No it does not signal. Target pathogen is bacteria. Ligands are LPS and proteins.

What is a common ligand among scavenger receptors?

LPS

Do TLRs induce phagocytosis?

No they dont. They only produce cytokine (signal).

What is the first thing that triggers an inflammatory response?

Pattern Recognition Receptors (PRRs)

What are PRRs?

They are molecular sensors that recognize structural motifs (pathogen molecular patterns (PAMPs)) in microbes that are highly conserved and usually necessary for their survival and innate and inflammatory responses.

Can PRRs also recognize DAMPs?

Yes

What are DAMPs?

They are damaged-associated molecular patterns. They are molecular patterns not normally seen in healthy cells and can trigger inflammation through PRRs.

What is an example of a DAMP?

Heat shock proteins are not expressed on healthy cells and will trigger TLRs or NLRs to cause the clearance of dead, dying, and aging cells by macrophage-mediated phagocytosis

Is RIG-I a PRR?

Yes it is

What cells are PRRs found on?

1. Myeloid cells

2. Lymphocyte subsets

3. Skin and mucosal epithelial cells (commonly exposed to pathogens)

4. Endothelial cells

5. Fibroblasts

What are the three types of PRRs?

1. TLR

2. NLR

3. CLR

What are TLRs? How do they bind?

They can detect a variety of PAMPs and DAMPs. They bind them via leucine rich repeat domains that make up the extracellular ligand-binding structure

What does ligand binding via TLRs induce?

TLR dimerization and signal transduction

What are NLRs?

They are an intracellular PRR that are activated by PAMPs and DAMPs

What are CLRs?

They bind carbohydrates on the surface of pathogens and promote phagocytosis