ST-1

• Common intracellular signal pathways via plasma membrane receptors

  • Receptors associated with G-proteins (GPCRs)
  • Receptors with protein kinase activity or other enzyme activity
  • Receptors associated with adapter proteins
  • Receptors associated with ion channels

  
• Plasma membrane receptors


  • Associated with ion channels
  • Associated with G-proteins (guanine nucleotide-binding proteins)
  • Receptors with intracellular domains with enzyme activity:

  1. Receptors with tyrosine kinase activity

  2. Receptors, associated with other intracellular enzymes

  1. with tyrosine kinases
  2. with other activity

  3. Receptors with tyrosine phosphatase activity

  4. Receptors with serine/threonine kinase activity

  5. Receptors with guanylate cyclase activity

  • Death receptors, triggering apoptosis
  • Integrins, involved in signaling associated with cell adhesion

  
• Receptors associated with G-proteins (GPCRs)

  • Receptors associated with ion channels
  • Receptors associated with other intracellular enzymes with tyrosine kinase activity
  • Receptors with intrinsic enzymatic activity (guanylate cyclase, tyrosine kinase)

  
• Nicotinic acetylcholine receptors (N-receptors)

  Ionotropic receptors - associated with ionic channels
• Binding of acetylcholine to Nicotinic acetylcholine receptors (N-receptors)

  1. Opening of Na+ and/or Ca2+ channels

  2. Depolarization of the membrane

  3. Excitation

  
• 1. proteins


  Guanine nucleotide-binding proteins

  
• Mechanism of action of G-proteins


  1. Activation/deactivation cycle dependent on binding to GTP or GDP

  2. Activation catalyzed by guanine nucleotide exchange factor (GEF)

  3. Deactivation catalyzed by protein that stimulates GTPase activity (GAP)

  
• Classes of G-Proteins


  • Gsα
  • Giα
  • Gqα
  • Gtα

  
• Gsα


  Activates adenylyl cyclase, increases cAMP
• Giα


  • Inhibits adenylyl cyclase, decreases cAMP
  • Activates K+ channel
• Gqα


  Activates phospholipase C, increases IP3 and DAG

  
• β-Adrenergic receptor activation


  Increases cAMP via Gsα
• α2-Adrenergic receptor activation


  Decreases cAMP via Giα
• Muscarinic acetylcholine receptor activation

  Activates K+ channel via Giα
• Hormones acting via adenylyl cyclase

  Increase cAMP (CRH, LH, hCG, LPH, PTH)
• Muscarinic acetylcholine receptor in heart muscle

  • Linked via G protein to K+ channels
  • Binding of acetylcholine activates Giα subunit
  • Gbsubunit binds and opens K+ channel
  • Increased K+ permeability hyperpolarizes membrane, reduces heart rate
• Receptor Tyrosine Kinases (RTKs)

  • Contain extracellular ligand binding domain, intracellular tyrosine kinase domain, intracellular regulatory domain, transmembrane domain
  • Classified into families based on extracellular structural features

  
• Steps in RTK signaling

  1. Ligand binding

  2. Receptor dimerization

  3. Autophosphorylation of tyrosine residues

  4. Binding of SH2 domain-containing proteins

  5. Activation of downstream signaling pathways

  
• SH2 domain


  Allows proteins to dock to phosphorylated tyrosine residues on other proteins
• Grb2


  Adaptor protein that binds phosphotyrosine residues on RTKs

  
• SOS


  Guanine nucleotide exchange factor (GEF) for Ras
• RTK signal transduction via Ras and MAPK pathway


  1. Grb2 binds phosphotyrosine on RTK

  2. Grb2 binds and activates SOS

  3. SOS activates Ras by promoting GTP binding

  4. Activated Ras binds and activates Raf (MAPKKK)

  5. Raf initiates MAPK phosphorylation cascade

  
• Small G proteins


  • Monomeric GTP/GDP binding proteins
  • Ras, Rho, Rab, Ran, Arf subfamilies
  • Activity depends on GTP/GDP binding state
  • Activation by GEFs, deactivation by GAPs

  
• MAP kinases


  Ser/Thr protein kinases involved in regulating cell processes

  Activated in MAPK cascades with 3 kinases: MAPKKK, MAPKK, MAPK

  
• Insulin receptor signaling


  1. Insulin receptor has intrinsic tyrosine kinase activity

  2. Phosphorylates IRS-1 adapter protein

  3. IRS-1 binds and activates PI3K

  4. Grb2 also binds phosphotyrosines on IRS-1
• Phosphatidylinositol 3-kinases (PI3K)


  Activated by tyrosine phosphorylation, regulates diverse cellular processes

  
• Insulin receptor (IR)


  Has intrinsic tyrosine kinase activity but does not directly interact (following autophosphorylation) with enzymatically active proteins containing SH2 domains (e.g. PI-3K or PLC-)
• IRS-1


  The principal IR substrate, contains several motifs that resemble SH2 binding consensus sites for binding the catalytically active subunit of PI-3K
• Insulin receptor signaling pathway

  1. IRS-1 acts as a docking or adapter protein to couple the IR to SH2 containing signaling proteins

  2. Additional adapter proteins have been identified, the most commonly occurring being a protein termed growth factor receptor-binding protein 2, Grb2

  
• Phosphatidylinositol 3-kinases (PI3K)

  • Activated by tyrosine phosporylation (by different RTKs) and it is associated with that receptors
  • A heterodymer: one of the subunit contains SH2 domain, which binds to activated receptors or other adaptor proteins, and allow the phosporylation and activation of the second catalityc subunit of PI-3K
  • Associated and activated by receptors of PDGF, EGF, insulin, IGF-1, HGF and NGF
  • Phosphorylates different phosphatidylinositols at position 3 in the inositol ring, creating additional substrates of PLC (stimulating a cascades mediated by DAG and IP3)
  • The obtained phosphatydilinositol-3,4,5-triposphate (PIP3) sequesters and activates a protein kinase, called PDK1, which on turn binds, posphorylates and activates a protein kinase B, called also AKT
  • AKT (PKB) regulates different cell responses
  • PTEN is a phosphatase, which dephosphorylates PIP3 and thus inhibits the signaling pathway

  
• mTOR (Mammalian target of rapamycin)

  • Serine/threonine kinase that regulates cell growth, cell proliferation, cell motility, cell survival, protein synthesis and transcription
  • Also know as FK506 binding protein 12-rapamycin associated protein 1 (FRAP1), coded by FRAP1 gene
  • Rapamycin is a bacterial toxin that inhibits mTOR by binding to its intracellular receptor FKBP12
  • Integrates the signal pathways of insulin, growth factors as IGF-1 and IGF-2, and some mitogens
  • May be consider as a sensor of cell nutrition and calorie demands, as well as of cell redox state
  • Some of the important target of mTOR are p70-S6 Kinase 1 (p70S6K1) and 4E-BP1 (eukaryotic initiation factor 4E (eIF4E) binding protein I)

  
• Non-Receptor Tyrosine Kinases (PTKs)

  • Numerous intracellular PTKs that are responsible for phosphorylating a variety of intracellular proteins on tyrosine residues following activation of cellular growth and proliferation signals
  • Two distinct families: one related to the Src protein, the second related to the Janus kinase (Jak)
  • Most of the proteins of both families of non-receptor PTKs couple to cellular receptors that lack enzymatic activity themselves, including all of the cytokine receptors (e.g. the interleukin-2 (IL-2) receptor) as well as the CD4 and CD8 cell surface glycoproteins of T cells and the T cell antigen receptor (TCR)

  
• Signal Transduction by JAK-STAT Receptors

  1. Tyrosine kinase-associated receptors called Jak-STAT receptors are often used by cytokines to regulate the proliferation of certain cells involved in the immune response

  2. The receptor itself has no intrinsic kinase activity but binds (associates with) the tyrosine kinase Jak (Janus Kinase)

  3. Their signal transducer proteins, called STATs (signal transducer and activator of transcription), are themselves gene-specific transcription factors

  4. There are 4 members of the JAK family (JAK 1,2,3 и Tyk2), two Jak kinases should be bound to the receptors

  5. The substrates of the JAK kinases are different members of STAT family

  6. STATs are inactive in the cytoplasm until they bind to the receptor complex, where they are also phosphorylated by the bound JAK

  7. Phosphorylation changes the conformation of the STAT, causing it to dissociate from the receptor and dimerize with another phosphorylated STAT, thereby forming an activated transcription factor

  8. The STAT dimer translocates to the nucleus and binds to a response element on DNA, thereby regulating gene transcription

  9. Jak-STAT receptors have a more direct route for propagation of the signal to the nucleus than tyrosine kinase receptors