13-cytokinesI, Microbiology Presentation

7/23/2019 13-cytokinesI, Microbiology Presentation

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INTERFERONs and CHEMOKINEs

CYTOKINE RECEPTORs/SIGNALINGADHESION MOLECULEs

Dr. Klára Megyeri

University of Szeged,

Faculty of Medicine,

Department of Medical Microbiology and Immunobiology

2014/2015

Type III IFNsType II IFNsType I IFNs

IFN-ζ (limitin)***

IFN-τ**

IFN-δ*IFN-κ

IFN-ε

IFN- λ3/IL-28BIFN-ω

IFN- λ2/IL-28AIFN-β

IFN-λ1/IL-29IFN-γ IFN-α

*Found in pig; **Found in ruminants; ***Found in mice

Classification of interferons (IFNs)

Production and action of IFNs

- -

Mechanism of IFN induction/production

PTK=Protein tyrosine kinase; TRIF=TLR adaptor molecule 1; TRAF=Tumor necrosis factor receptor-associated factor; TBK1=TRAF family member-associated NF-kappaB activator (TANK)-binding kinase 1; IRF=Interferon regulatory factor;TAK1=Transforminggrowth factor-β-activatedkinase 1; RIG-I=Retinoic acid-inducible geneI; MDA 5= Melanoma differentiationassociated protein 5; IPS1=IFN-β-promoter stimulator 1; AIP3=Atrophin-1 interacting protein 3; CCL5=Chemokine (C-C motif)ligand 5; CXCL10=Chemokine (C-X-C motif) ligand 10; IFIT1/2=Interferon-induced protein with tetratricopeptide repeats 1/2;IKK=Inhibitor of NFκ B kinase; SELE=Selectin E (endothelial adhesionmolecule1)



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Endosomal or cytoplasmic double-stranded (ds) RNA produced during viralreplication can activate different signaling pathways triggered by eithermembrane-bound TLR3 or cytoplasmic RIG-I or MDA5.a | TLR3-dependent pathway

TLR3 recognizes dsRNA in the lumen of the endosome, which causes phosphorylation of specific tyrosine residues in TLR3 by an unidentified protein tyrosine kinase.

TLR3 dimerizes, binds to CD14 and activates the signalling complex assembled by TRIF.Two major pathways bifurcate from TRIF:•One, composed of TRAF3 and TBK1/IKKE, leads to phosphorylation of the transcriptionfactor IRF3.•The other branch acts through TRAF6 and TAK1 leading to the activation of NFκ B, JUNand ATF2 transcription factors.

The activated transcription factors translocate from the cytoplasm to the nucleus, bindto the cognate sites in the promoters of the target genes and singly or in combinationsinduce their transcription.

b | RIG-I/MDA5-dependent pathway

The cytoplasmic RNA helicases RIG-I and MDA5 recognize dsRNA or 5′ triphosphorylatedsingle-stranded (ss) RNA and use the mitochondrial membrane-bound protein IPS1 as thespecific adaptor.

IPS1 functions like TRIF and activates the same transcription factors leading to theinduction of similar genes.

In addition, they cause apoptosis by activating caspases 8 and 10 through the interactionof FADD withIPS1.

IFNs interact with IFN receptors (IFNAR1, IFNAR2 or IFNLR1). Thesereceptorsare associated with two kinases fromthe JAK family: JAK1 and TYK2 for type I and III IFNs; JAK1 andJAK2 for type II IFN.

IFN-binding triggers a JAK/STAT signaling cascade consisting of two Janus tyrosine kinases (JAK-1 and TYK-2) andtwo members of signal transducers and activators of transcription (STAT-1 and STAT-2).

Phosphorylated form of STAT-1 dimerize (signaling activated by Type II IFNs) or phosphorylated STAT-1/STAT-2heterodimers associate with a p48 protein (IRF-9) constituting a transcriptional activation complex ISGF-3 (signalingactivated by TypeI andType III IFNs).

Activated STATs translocate to cell nucleus and bind to IFN-stimulated response elements (ISRE) or IFN-γ γγ γ -acivatedsites (GAS) within promoter regions of IFN-inducible genes.

IFN receptors and signaling

There is a large number of IFN-inducible genes (>300), including

PKR, 2’5’-OAS, Mx, ADAR, IRFs and MHC

The IFN-inducible genes encode proteins that mediate the antiviral,

antiproliferative and the immunomodulatory effect of IFNs.

The molecular mechanism of IFN-mediated antiviral effect



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OAS/RNaseL

The antiviral and anti-tumor actions of interferons are caused, in part, by a remarkableregulated RNA cleavage pathway known as the 2-5A/RNase L system.

Elimination of virus-

infected cell by apoptosis5. rRNA and other

Production of small duplex

RNAs that induce IFNs

4. Self RNA (host)and non-self RNA

(virus)

Damage to host cell

machinery required for viral

replication

3. Cellular mRNA

and rRNA

RNase L

Inhibition of viral protein

synthesis

2. Viral mRNA

(any virus)Active

Elimination of viral genome

1. Viral genomic

RNA (ssRNA

viruses only)

Antiviral effectRNA substrates

(2'-5')(A)n synthetase=OAS/RNase L enzyme systemType: multi-enzyme systemMechanism of action:The (2'-5')(A)n synthetase=OAS adds 2',5'-oligoadenylate moiety to ssRNA molecules → Theintracellular appearance of 2',5'-oligoadenylated RNA leads to the activation of RNaseL,which in turncleaves the 2',5'-oligoadenylated RNAs → Degradation of viral ssRNAmolecules blocks replication of viral genomic nucleic acid, and inhibits viral protein synthesis

PKR=dsRNA-dependent protein kinaseType: serine-threonine kinaseMechanism of action:IFN increases the level of PKR, and viral dsRNA activates it → PKR inactivates eIF2α(eukaryotic Initation Factor 2α) by phosphorylation → inactivation of eIF2α blocks cellular

translation and synthesis of viral proteins

Mx proteinsType: GTPase (dynamin family)Mechanism of action:Mx proteins inhibit transcription of viral genes therebyblock the multiplication of viruses

ADAR1=Adenosine deaminase acting on RNAType: RNA editing enzymeMechanism of action: The enzymatic activity (deamination of adenosines) of ADAR resultsin A(adenosine)-to-I(inosine) transition that affects the stabilityof duplex RNA caused by aconversion of stable AU base pairs into a less stable IU. Adenosine deaminase may cause ahypermutability by extensive A-to-I editingto a number of viruses (including measles,parainfluenza type 3, vesicular stomatitis and polyomavirus) that eventually may lead topersistent infection.

1.

2.3.

4.

1. Activates macrophages

2. Regulates isotype switching

3. Promotes TH1 differentiation

4. Increases MHC expression and

antigen presentation

The major immunomodulatory effects of Type II IFN=IFN-γ γγ γ



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The major biological effects of type III IFNs

(A) Targets of type III IFNs mainly include epithelial cells from the skin, the gut, the respiratory

system, andthe urogenital system, as well as hepatocytes.

(B) On their targetcells, typeIII IFNs exert antiviral(top) and antibacterial (bottom) actions.

(C) Whether the antiproliferative of type III IFNs is relevant for natural antitumor defense must be

clarified in future investigations. CGD (chronic granulomatous disease; mutation in the NADPHoxidase gene)

IFN-γ γγ γ Imukin

Malignant melanomanatural IFN-ααααMultiferon

Multiple SclerosisIFN-ββββ1bExtavia

Multiple SclerosisIFN-ββββ1aRebif

Multiple SclerosisIFN-ββββ1aAvonex

Multiple sclerosisIFN-ββββ1bBetaferon

Chronic hepatitis C, hepatitis BPegylated IFN-αααα2aPEGASYS

Chronic hepatitis CPegylated IFN-αααα2bPEG-Intron

Condyloma acuminatumIFN-ααααn3Alferon

Chronic hepatitis CIFN-ααααcon1Infergen

Chronic myeloid leukemia, hairy cell leukemia, AIDS-related Kaposi’ssarcoma, chronic hepatitis C, hepatitis B, renal carcinoma, follicular non-Hodgkin lymphoma, malignant melanoma, cutan T-cell lymphoma

IFN-αααα2aRoferon-A

Hairy cell leukemia, condyloma acuminatum, follicular lymphoma, malignantmelanoma, AIDS-related Kaposi’s sarcoma, chronic hepatitis B, hepatitis C

IFN-αααα2bIntron-A

IndicationsCytokineProduct

IFN therapy

Chemokines are small Mw (8-10 kDa) secreted polypeptides that are chemotactic for

cells in the immune system such as leukocytes which move up the gradient of

chemokine secreted by another cell; thus, they control the temporal and spatial

positioning of leukocytes during an immune response. Based on their structure,

chemokines can be classified into 4 groups:

Producer cells: monocytes, endothelial cells, keratinocytes,

fibroblasts, smooth muscle cells, T-cells

CHEMOKINESCHEMOKINES

α-chemokines (CXC): Act primarilyon

the neutrophil granulocytes

β-chemokines (CC):

Actprimarily onthe monocytes

γ -chemokines (C): Actprimarily onthe

T-cells

δ-chemokin (CX3C)



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Combinatorial interactions of the chemokines and their receptors Certain viruses (e.g.: HIV) may infect cells by binding to chemokine receptors

The functions of chemokines

Chemokines play important roles in several physiological and pathological

processes:

Physiological role:

•in the chemotactic movement and adhesion of leukocytes

•in innate and adaptive immune responses

•in cell proliferation

•in cell differentiation

•in apoptosis

•in angiogenesis

•in hematopoiesis

•in ontogeny

Pathological role:

•in inflammation, infections

•in the formation of malignant tumors (in cell transformation and dissemination)

multiple sclerosis, rheumatoid arthritis, diabetes, endometriosis, transplant rejection,

atopic dermatitis, psoriasis, ulcerative colitis, Crohn-disease, ischemia,

atherosclerosis, multiple myeloma

Cytokine-receptor families

1. Type I cytokine-receptor family (Hemopoietin cytokine-receptor family): adomain with two conserved pairs of cystein residues and tryptophan-serine-X-tryptophan-serine (WSXWS) sequence

2. Type II cytokine-receptor family: a domain with two conserved pairs of cysteinresidues, without WSXWS sequence

3. Tumor necrosis factor (TNF)-receptor family: cysteine-rich extracellulardomain

4. Immunglobulin superfamily receptors: immunoglobulin extracellular domain

5. Seven transmembrane αααα-helical receptors (Serpentine receptors): theirtransmembrane domains appear to ”snake” back and forth through the membrane





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Negative regulation of the Jak-Stat

signal transduction pathway mayinhibit the overstimulation of cells.

SOCS (8)= Suppressor of cytokine signalling (inhibit phosphorylation and

activation of the receptor, Jak and Stat by different mechanisms; classic feedback

inhibitors)

PTP= Protein tyrosine phosphatases (by way of dephosphorylation inactivate the

receptor, Jak and Stat molecules)

PIAS= Protein inhibitor of activated Stat (inhibit the transcriptional activity of Stat

molecules)

TNFTRAF-mediated

TNFR pathway:

TRADD

TRAF

RIP

SAP kinase cascade

↓↓↓↓JNK

TRANSCRIPTION OF TNF-RESPONSIVE GENES:

PRODUCTION OF INFLAMMATORY MEDIATORS AND

SURVIVAL PROTEINS

c-jun c-fos

AP-1 Iκ κκ κ B

NF-κ κκ κ B

NF-κ κκ κ B Iκ κκ κ B

Iκ κκ κ B kinase (IKK) cascade

Death domain-

mediated TNFR

pathway:

TNF

TRADD

FADD

Inactive caspase-8

Active caspase-8

Activation of the effector caspases

APOPTOSISAPOPTOSIS

•Degradation of the structural proteins

•Deregulation of the cell cycle progression

•Inhibition of DNA replication and repair

•Modulation of the cellular signal transduction

•Activation of certain cytokines

Degradation of the nuclear

matrix

INTERNUCLEOSOMAL

DNA FRAGMENTATION

CHEMOKINE

Gαβγ

GDP

βγ Gα GTP

Adenylate cyclase↓↓↓↓

cAMP↓↓↓↓

PKA

↓↓↓↓

CREB

PLCββββ↓↓↓↓

PIP2

IP3↓↓↓↓

Ca2+

↓↓↓↓Calcineurin

↓↓↓↓NFAT

↓↓↓↓

Iκ κκ κ B

NF-κ κκ κ BIκ κκ κ B

DAG↓↓↓↓

PKC↓↓↓↓

NF-κ κκ κ B

JNK/ERK

↓↓↓↓

c-myc

c-fos c-jun

G protein

pathway:

TRANSCRIPTION OF CYTOKINE-RESPONSIVE GENES







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Inside out signaling

kindlin

Leukocyte adhesion deficiency-I-III (LAD)

LAD-I: type 1 leukocyte adhesion deficiency= a CD18 gene (ββββ2-integrin; ITGB2 gén) mutation,

leads to recurrent bacterial and ungal infections, PMN cell iniltration is missing in the tissues,impairment of adherence-dependent leukocyte functions

LAD-II: fucose-transporter gene mutation, leads to decreased expression of Lewis X carbohydrate.Thus, the ligand of E- and P-selectin is missing from the surface of leukocytes. Clinically similar toLAD-1.

LAD-III: mutation in the gene encoding kindlin-3 cytoplasmic protein (FERMT3). This protein isessential for the activation of integrins (implicated in the „inside out signaling” process). Adhesion of leukocytes and platelets is decreased. Clinically similar to LAD-I+ Glanzmannthrombasthenia.

LAD-2 LAD-3 LAD-1

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13-cytokinesI, Microbiology Presentation