Immunology and Infection

Physical Barriers

Intrinsic mechanisms

=> Unspecific, immediate

Skin

Saliva

Mucus

Tears

Defensins

RNAi (RNA Interference)

Epigenetic Silencing

Apoptosis

Innate (after minutes to hours)

Adaptive (after hours to days)

NK (Natural Killer) cells

Cytokines, Interferons (alpha, beta)

Complement system (part of the humoral innate system, enhances (= complements) the ability of antibodies and phagocytic cells to clear microbes and damaged cells from an organism, promote inflammation, and attack the pathogen's cell membrane)

Sentinel (Mast cells, Dendritic cells, macrophages)

Cell mediated (Interferon gamma, CTL = cytotoxic T Lymphocytes, NK cells and macrophages)

Humoral (antibodies)

HIV

Hepatitis B & C

siRNA: Plants, Insects, Fungi

miRNA: Mammals (, vertebrates?)

For miRNA:

1. In Nucleus: double stranded Pri-miRNA is cut into pre-miRNA via Drosha

2. pre-miRNA gets transported into the cytoplasm via Exportin 5 and cut into miRNA by DICER

3. miRNA double strands are separated via RISC/AGO complex and passenger strand is discarded

4. RISC-enzyme complex gets activated by miRNA and leads to incomplete complementary binding of the guide strand to target mRNA (virus mRNA)

5. Binding leads to translational repression (in mammals), mRNA degradation and mRNA cleavage (not in mammals)

For siRNA:

1. DICER cuts viral dsRNA into double stranded siRNA

2. RISC/AGO enzyme complex cleaves the passenger strand and gets activated

3. Complementary Binding of the guide strand to target mRNA (virus)

4. mRNA gets cleaved

Cycle: Restriction of virus through host and countermeasures of the virus against restriction

Viral miRNAs are 100% complementary to other viral miRNA => get cleaved => feedback loop by the virus to stay under the radar

Hallmarks:

Chromatin Condensation

DNA Fragmentation

Apoptotic Bodies

Plasma Membrane Integrity (membrane is stable during the whole process)

Induction Pathways:

Intrinsic = mitochondrial pathway

Extrinsic = Death receptor pathway

Hepatitis B: Inhibition of p53 (transcription factor inducing apoptosis if there is DNA damage) dependent apoptosis via protein HBx

Polyomaviruses/Papillomaviruses: Degradation/Inhibition of p53 via protein LT-antigen

Humanes Herpesviruses: Inhibition of caspase cascade (get activated during cell stress (intrinsic) or by DISC (extrinsic)) via protein v-FLIP/K13 (FLICE inhibitor)

Adenovirus, EBV: Bcl2 Homologue (protein E1B 19K), TNF alpha inhibition (protein LMP-1)

APOBEC3G belongs to the family of cytidine deaminases that catalyze the deamination of cytidine to uridine in the single stranded DNA substrate

Is expressed within non-permissive cells (resting T cells, macrophages)

Apobec 3G deaminates C (Cytosin) to U (Uracil) in DNA = non-sense transcription = HIV non-infectious in the next cell

Countermeasures of HIV resrticts Apobec 3G via Vif: ubiquinates A3G via Vif Enzyme complex (Cul5, Ned8, EloB-EloC, Cbf-beta, E2, Rbx-2) and leads to degradation in the proteasome = HIV infectious again

Vif1 binds to A3G as well as the cellular Cullin5 E3 Ubiquitin Ligase (ELOB-ELOC-CUL5) and a CBFB cofactor so that the ligase can be hijacked to tag A3G for degradation

Function/Principle: Condensing the chromatin of viral genomes via histone marks to repress transcription

Activating histone marks: H3K27Ac, H3K4me3

Repressive histone marks: H3K27me3, H3K9me3

3 pathways

1. PML-ND10 nuclear bodies: Membraneless nuclear structures Includes: PML (promyelocytic leukaemia protein), Sp100, Smc5, Smc6, Daxx and ATRX Function: Deposits histones with repressive marks (histone H3.3)

2. IFI16 (Interferon gamma Inducible Protein 16): Innate Immune sensor for intracellular DNA Function: Deposition of repressive histone methylation

3. NP220 + HUSH (human silencing hub) complex: Deposit heterochromatic marks on unintegrated viral DNA (specifically for the retrovirus murine leukaemia virus)

Herpes simplex virus 1 (HSV-1): Protein VP16 associates with the host factors HCF-1 and Oct-1, with HCF-1 recruiting the host H3K9 demethylases LSD1 and JMJD2 to remove repressive marks, along with the H3K4 methyltransferases Set1 and MLL1 to deposit active marks on viral DNA

Several viral proteins target pro-myelocytic leukaemia nuclear body (PML-NB) components in order to avoid epigenetic repression.

=> HSV-1 ICP0 and human cytomegalovirus (HCMV) IE1 induce the degradation of PML and Sp100

=> Epstein–Barr virus (EBV) BNRF1 disassembles the Daxx–ATRX histone chaperone complex, whereas HCMV pp71 degrades Daxx

=> Hepatitis B virus (HBV) protein HBx induces the degradation of Smc5 or Smc6.

HSV-1 can also induce the degradation of IFI16 with ICP0 implicated as being involved in this degradation.

Retroviruses avoid epigenetic repression by integrating their DNA into host euchromatin, where it can no longer be identified as foreign DNA. MLV, murine leukaemia virus

= Pathogen associated molecular pattern

Lead to expression of IFN-alpha/beta => activate ISGs (interferon stimulated genes) = alpha-viral effector proteins (PKR,Mx, APOBEC, Trim5alpha, 2’5’OAS => RNAseL, Tetherin)

PAMP-Sensoren:

1. TLRs: TLR7,-8 (ssRNA), TLR3 (dsRNA), TLR9 (unmethylated DNA)

2. RLRs: RIG-I (ssRNA), MDA5 (dsRNA), cGAS/STING (cytosolic DNA)

Type I Interferons (IFN- α and IFN-β; 13 IFN- α subspecies, 1 single IFN- β gene, IFN-ε; IFN- κ + IFN- ω (single gene, produced by epithelial of specific tissues)

Direct response to viral infection

pDC and macrophages produce high levels of IFN- α

Fibroblasts mainly produce IFN-β

Tightly regulated on transcriptional level (promoter of IFN-β contains binding sites (ISRE) for IRF3, IRF7 and NFkB)

IFN- α subtypes bind to type I IFN receptor (2 subunits) => Heterodimerization, conformational changes, activation of signalling cascade

> 1000 genes are activated by stimulation of type I IFNAR

==> Antiviral, anti-proliferative, immunomodulatory functions

Function: cell to cell interaction and signalling + host antimicrobial defence (vertebrates) ==> NF-kB activation, MAPK activation, Expression of pro-inflammatory cytokines, Maturation of DC

Extracellular domain: LRR = leucine rich repeat (22-29aa repeats; protein-protein interaction; binding to carbohydrates, lipids and nucleic acids)

=> pathogen recognition

Intracellular domain: TIR = Toll/Interleukin 1 receptor; Five TIR-containing cytosolic adaptors (MyD88, TIRAP, TRIF, TRAM, SARM)

=> downstream signalling

TLR1,2,4,6: recognize lipids

TLR3,7,8,9: recognize nucleic acids

TLR1,2,4,5,6,10 are expressed on cell surface

TLR3,7,8,9 are expressed in intracellular compartments (endosome)

At least 13 TLR members in mammals

= plasmacytoid DCs

recognize viral nucleic acids

N-terminal DD (Death domain); C-terminal TIR (Toll/Interleukin receptor) domain

TLR7,8

ssRNA

IRAK-4 (Interleukin 1 R associated kinase) binds to MyD88

TIR Domain containing adaptor IFNβ

TLR3

dsRNA

Detect PAMPs

RIG-I

MDA5

cGAS/STING

RIG-1 (Retinoic acid inducible gene I): DExD/F box RNA helicase domain

MDA-5 (melanoma differentiation associated protein 5): DExD/F box RNA helicase domain

MAVS (mitochondrial antiviral signalling proteins): IPS1 (Interferon promotor stimulating Factor I), CARD-like domain

cGAS: binds to dsDNA/cytoplasm => undergoes conformational change to build cGAMP (cyclic dinucleotide)

STING: stimulator of Interferon genes

TBK1: Tank-binding kinase 1

Recognized by RIG-1: Paramyxoviruses (Sendai virus, parainfluenza), Orthomyxoviruses (Influenza), Rhabdoviruses (vesicular stomatitis virus (VSV)), Flaviviruses (Hepatitis C virus (HCV), Japanese encephalitis virus)

Recognized by MDA-5: Picornaviruses (Rhinovirus, Theiler’s disease virus)

Recognized by both: Flaviviruses (West Nile virus (WNV), Dengue virus)

an ISG

Expressed at low levels in all cells

Induced by IFN (autophosphorylation, homodimerization)

Activated by cellular and viral RNA with partial ds structures, heparin and other polyanionic compounds

Translation inhibition and apoptosis as result of PKR activation

Overexpression of PKR = inhibition of HBV replication

Viruses inhibit PKR by either sequestering PKR through viral binding (Adenovirus or HIV) or inhibiting PKR phosphorylation (HepC Virus NS5A protein)

an ISG

Type II integral membrane protein

IFN induced expression in other cells

constitutively expressed in mature B cells, plasma cells and plasmacytoid dendritic cells

inhibits release of many enveloped viruses (HIV, EBOLA, LASSA, MARBURG Virus, CMV)

Viruses counteract by targeting Tetherin to proteasomal degradation (HIV) or lysosomal degradation (KSHV)