CYTOKINE RECEPTORS AND SIGNAL TRANSDUCTION

Survival factor (eg. IGF-1) ChemokinesHormonesTransmitters (e.g. Interleukins serotonin etc.) Growth factors (eg TGFa) Extracellularmatrix Wnt Hedgehog Death factors (eg TNF) Cytokines

What is Cytokine?  Secreted polypeptide or low molecular weight protein involved in cell-to-cell signaling.  Acts in paracrine or autocrine fashion through specific cellular receptors.  Can be produced by cells of any tissue and act on many cells involved in immune and inflammatory response.

Cytokines, like hormones, can act in autocrine, paracrine, or endocrine fashion

Cytokine nomenclature  Lymfokines  Lymfokines - produced by activated T lymphocytes direct the immune system response by signaling between its cells  Interleukins -  Interleukins - presumed targets are principally leukocytes.  Chemokines  Chemokines - specific class of cytokines. Mediates chemoattraction (chemotaxis) between cells, stimulate leukocyte movement and regulate the migration of leukocytes from the blood to tissues.  Monokines  Monokines - derived primarily from mononuclear cells such as macrophages.

Cytokines: main functions  Hematopoiesis  Hematopoiesis (ex. CSFs, colony stimulating factors).  Inflammatory reaction  Inflammatory reaction (ex. IL1, TNF).  Chemotaxis  Chemotaxis (ex. IL8, MIP1- macrophage inflammatory protein 1, BLC – B-lymphocyte chemoatractant ).  Immunostimulation  Immunostimulation (ex. IL12, IFN  ).  Suppression  Suppression (ex. IL10).  Angiogenesis  Angiogenesis (ex. VEGFs - vacsular endothelial growth factor ).  Embryogenesis  Embryogenesis (ex. TGF- , LT - lymphotoxin ).

 Type I cytokine receptor or hematopoietin receptor family : IL 2, IL 3, IL 4, IL 5, IL 6, IL7, IL 9, IL 13, IL 15, GM- CSF (Granulocyte-Macrophage Colony-Stimulating Factor) and G-CSF (Granulocyte-Colony Stimulation Factor)  Type II cytokine receptors or interferon receptors  Transforming growth factor receptor  Tumor necrosis factor receptor  Immunoglobulin superfamily (IgSF)  Chemokine receptors (seven transmembrane helix) Classification

Type I cytokine receptor Transmembrane receptors expressed on the surface of cells. These receptors are also known under the name hematopoetin receptors, and share a Trp-Ser-X-Ser- Trp motif (WSXWS) in the extracellular portion adjacent to the cell membrane.

Common  chain The  chain (green), common to all, mediates intracellular signaling.

Signaling through cytokine receptor Phosphorylation through kinases: The addition of a phosphate molecule to a polar R- group of Tyr can turn a hydrophobic portion of protein into a polar and extreme hydrophilic portion of molecule. – Kinase is a type of enzyme that transfers phosphate group (PO 4 ) from high-energy donor molecules, such as ATP to specific target molecules (substrates). – The opposite, an enzyme that removes phosphate groups from targets, is known as a phosphatase. – Kinase enzymes that specifically phosphorylate tyrosine amino acids are termed tyrosine kinases.

Signaling through cytokine receptor The ligand-activated receptor (R) attracts a Janus (JAK) kinase (K). K phosphorylates both itself and the receptor. A Signal Transducer and Activator of Transkription (STAT) protein (S) binds to tyrosine-phosphorylated receptor-kinase complex. After being phosphorylated by JAK, the STATs form active dimers that translocate into the nucleus to regulate transkription.

  Protein hormones with antiviral activity.   Secrete by cells in a response to variety of stimuli.   Type I and type II IFN and IFN-like cytokines.   Effects are mediated through cell receptors.   IFN activate cellular signalling pathway (gene induction or repression). Type II cytokine receptors Interferons (IFN)

Type I IFNs consist of seven classes: IFN- , IFN- , IFN- , IFN- , IFN-  IFN-  and IFN-    Type I IFNs are major components of the innate immune system.   Protect against viral infection.   The expression of type I IFNs is induced by viral challenge. Type II IFN consist of IFN-  only.   IFN-  (immune interferon) is produced by certain activated T-cells and NK cells.   IFN-  is made in response to antigen (including viral antigens) or mitogen stimulation of lymphocytes.

Type I IFNs   Produced by macrophages, neutrophils and other somatic cells in response to infection by viruses or bacteria.   Inducer is double strand RNA provided by viral genom itself.   Receptors are expressed on most cell types. IFN-    IFN-  is produced in activated T H 1 and NK cells, particularly in response to IL-2 and IL-12.   Binding of IFN-  to its receptor increases the expression of class I MHC on all somatic cells.   IFN-  may also activate macrophages, neutrophils and NK cells.

Initiation and regulation of variety responses   antiviral   antiproliferative activity (ability to arrest cell growth) – treatment for cancer   control of apoptosis   immunomodulatory (INF-  predominantly modulates immune response, main antiviral cytokine). Function of IFNs:

IFN- receptor   Expression of on the surface almost of all cell types.   High affinity receptors are located in the T- and B-lymphocytes, NK-cells, monocytes, macrophages, neutrophiles, fibroblasts, endotelial cells and smooth muscle cells.   Receptor is expressed only in response to stimulus by antigen, only in cells of lymphoid origin (NK cells, macrophages, and some T cells).

IFN-  R kDa glycoprotein (315 amino acid residues) Extracellular domain – 226 amino acid residues Transmembrane domain – 23 amino acid residues Intracellular domain – 65 amino acid residues. Ligand binds to extracellular domain of IFN -  R1 only (in absence of IFN -  R1, IFN -  R2 cannot bind IFN  IFN-  R kDa glycoprotein (472 amino acid residues) Extracellular domain – 228 amino acid residues Transmembrane domain – 24 amino acid residues Intracellular domain – 220 amino acid residues IFN  receptor has two components:  R1 and  R2

 Jak  Jak proteins are brought into close after ligand-receptor complex formation.  Jak1 Jak2Jak2 Jak1Jak1Jak2  Transphosphorylation between Jak1 and Jak2 proteins (Jak2 phosphorylates Jak1, Jak1 transphosphorylates Jak2).  Tyr 440  phosphorylation of IFN-  R1 (Tyr 440).  STAT1  Binding of STAT1 protein to each IFN-  R1.  S   Bound S  is phosphorylated by Jak.   Dissociation of dimer from the receptor and formed dimer translocates to the nucleus.   Induction of transcription of many genes. Signal transduction is carried out through a series of tyrosine phosphorylation events and culminates with the activation and nuclear translocation of STAT protein and new mRNA synthesis is induced.

IFN  receptor   IFN-  R amino acid residues (409 residues of protein are extracellular, 100 residues are intracellular.   IFN-  R AA residues in extracellular space, 251 AA residues in intracellular space.   Both components bind type-IFNs cooperatively.   Receptor has ability to bind multiple ligands (all subspecies of IFN-  and IFN-  and other types of IFN-type I).   Interferon ligand is boud to IFN-  R1 and than to IFN-  R2 which stabilizes the complex.

Tyk2 associates with IFN-  R1 Jak1, STAT1 and STAT2 associate with IFN-  R2.   Jak1 transphosphorylates Tyk2 (1). Tyk2 in turn phosphorylates Jak1 and IFN-  R1 (2).   Phosphorylation of IFN-  R1 allows STAT2 to bind to IFN-  R2.   STAT2 phoshorylates STAT1.   STAT1-STAT2 complex dissociates from receptor.   Dimer STAT1-STAT2 associates with interferon regulatory factor to form the transcription complex. Signal transduction goes through Jak/STAT pathway. Involves two different ligand binding proteins (kinases) Tyk2 and Jak1.

  Induces inflammatory reaction.   Induces antibacterial effect (activation of neutrophils, NK cells and macrophages, increased their ability to recognize, kill, and digest foreign materials or microbes).   Normal expression of is important in preventing the development of cancer. Effect of signalling through IFN-  receptor Effect of signalling through IFN-  /  receptor Antiviral defence (protects the cell from viral replication).

Immunoglobulin superfamily (IgSF)  The immunoglobulin superfamily (IgSF) - a large group of cell surface and soluble proteins that are involved in the recognition, binding, or adhesion processes of cells.  Molecules are categorized as members of this superfamily based on shared structural features with immunoglobulins (antibodies); they all possess a domain known as an immunoglobulin domain or fold.  Associated with roles in the immune system.

Tumor Necrosis Factor receptor Typical structure of a chemokine receptor, with seven transmembrane domains. Chemokine receptors are usually linked to a G-protein through which they signal. Interaction with their specific chemokine ligand, chemokine receptors trigger a flux in intracellular calcium (Ca2+) ions (calcium signaling) and cause cell response (chemotaxis). Cysteine-rich common extracellular binding domain Chemokine receptor

 After ligand binding, (PDGFR or EGFR receptor tyrosine kinases (RTKs) dimerize, undergo autophosphorylation (P) and recruit adaptor proteins (such as GRB2 and SHC) that activate various downstream effectors.  RAS is an important downstream effector and functions as a molecular switch by cycling between the active GTP-bound form and the inactive GDP- bound form. RAS activity is regulated positively by guanosine exchange factors (GEFs), such as SOS, and negatively by GTPase-activating proteins (GAPs).  At least three downstream effectors can be activated by RAS. The RAF- mediated signalling cascade. Transforming Growth Factor Receptor

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