Cell Signaling II Signal Transduction pathways

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Cell Signaling IISignal Transduction pathways

• Cell Biology
• Lecture 13

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Readings and Objectives

• Reading
• Russell Chapter 8 (not sufficient)
• Cooper Chapter 15
• Topics
• Lecture 12
• Signaling Molecules and Their Receptors
• Functions of Cell Surface Receptors
• Lecture 13
• Pathways of Intracellular Signal Transduction
• Signal Transduction and the Cytoskeleton
• Signaling Networks

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Intracellular Signal Transduction Pathways

• Intracellular signal transduction- chain of
  reactions, transmits signals/cell
  surface?intracellular targets
• First studied for epinephrine
• Signals glycogen breakdown to glucose
• Earl Sutherland (1958) action of epinephrine was
  mediated by an increase in cyclic AMP (cAMP)
• Concept cAMP is a second messenger
• Noble prize 1971

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cAMP Signal Transduction Pathways

• Epinephrine receptor coupled to adenylyl cyclase
  via a G protein? increasing the concentration of
  cAMP
• cAMP signaling cell responses
• Metabolic regulation
• Cytosolic Protein Kinase A activation (PKA)
• tetramer of regulatory and catalytic subunits, ie
  R2C2 (inactive)
• cAMP binding of R? dissociation of catalytic
  subunits (active)
• A serine/threonine kinase?Activation or
  inactivation of substrate proteins

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cAMP Signal Transduction Pathways

• Phosphorylation of two downstream enzymes
• Glycogen synthase inactivated? glycogen
  synthesis?
• Phosphorylase kinase activated? phosphorylates
  Glycogen phosphorylase (active)? Glu-1P?

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cAMP Signal Transduction Pathways

• 2. Gene regulation
• Increased cAMP activate transcription
• Free PKA C-subunit translocated to the nucleus
• binds Genes containing a regulatory sequencethe
  cAMP response element, or CRE
• phosphorylates the transcription factor CREB
  (CRE-binding protein).
• Recruits RNA polymerase
• expression of cAMP-inducible genes
• Proliferation, differentiation, memory,
  cognition
• Review article Transcriptional regulation by cAMP

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cAMP Signal Transduction Pathways

• Protein phosphorylation is reversed by protein
  phosphatases
• terminates responses initiated by receptor
  activation of protein kinase

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Secondary messenger DAG and IP3 signaling

• PLC-? binds receptor protein tyrosine kinases
  via SH2 domain? phosphorylated (active)
• PLC- ? stimulates hydrolysis of PIP2 to DAG and
  IP3 (how?)
• DAG and IP3 are secondary messengers
• IP3 regulates Ca2
• DAG activates PKC family

PLCPhospholipase C PIP2 Phosphatidylinositol
4,5-bisphosphate IP3 Inositol
1,4,5-trisphosphate DAG Diaceyl glycerol
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Secondary messenger DAG and IP3 signaling

• PLC-? binds receptor protein tyrosine kinases
  via SH2 domain? phosphorylated (active)
• PLC- ? stimulates hydrolysis of PIP2 to DAG and
  IP3 (how?)
• DAG and IP3 are secondary messengers
• IP3 regulates Ca2
• DAG activates PKC family

PLCPhospholipase C PIP2 Phosphatidylinositol
4,5-bisphosphate IP3 Inositol
1,4,5-trisphosphate DAG Diaceyl glycerol
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Secondary messenger DAG and IP3 signaling

• DAG remains associated with the plasma membrane
  and activates protein-serine/threonine kinases of
  the protein kinase C family.
• IP3 , a small polar molecule, released to the
  cytosol
• Stimulates release of Ca2 from the ER by binding
  to receptors that are ligand-gated Ca2 channels

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Secondary messenger DAG and IP3 signaling

• Calmodulin is activated when Ca2 concentration
  increases
• CaM kinase family are activated by
  Ca2/calmodulin
• they phosphorylate and activate other proteins
  such as,
• protein kinases, phosphatases, metabolic enzymes,
  ion channels, and transcription factors (eg CREB)
• Also regulates synthesis and release of
  neurotransmitters

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Secondary messenger DAG and IP3 signaling

• nonmuscle cells and smooth muscles, contraction
  is regulated by phosphorylation of myosin light
  chain
• catalyzed by myosin light chain kinase, which is
  regulated by the Ca2 binding protein calmodulin

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Secondary messenger DAG and IP3 signaling

• Increased Ca2 signals further release of Ca2
  from the ER by opening Ca2 channels (ryanodine
  receptors) in the ER membrane.
• Ca2 is a versatile second messenger that
  controls a wide range of cellular processes
• These pathways function coordinately to regulate
  many cellular responses

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PI 3/Akt signaling pathway

• PIP2 is also the start of another signaling
  pathway
• PIP2 is phosphorylated by phosphatidylinositide
  (PI) 3-kinase
• This yields a second messenger,
  phosphatidylinositol 3,4,5-trisphosphate (PIP3)

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PI3/Akt signaling pathway

• PIP3 targets a protein-serine/threonine kinase
  called Akt and also binds protein kinase PDK1
• Activation of Akt also requires protein kinase
  mTOR (in a complex called mTORC2) which is also
  stimulated by growth factor

mTOR mammalian target of rapamycin PDK1
phosphoinositide dependent protein kinase-1 GSK3
glycogen synthase kinase 3 Bad Bcl2 associated
death promoter (promotes apoptosis)
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PI3/Akt signaling pathway

• Akt phosphorylates several target proteins,
  transcription factors, and other protein kinases
• Transcription factors include members of the
  Forkhead or FOXO family
• If growth factors are not present, Akt is not
  active
• FOXO travels to the nucleus, stimulates
  transcription of genes that inhibit cell
  proliferation, or induce cell death

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PI3/Akt signaling pathway

• When growth factors attached to receptor/tyrosine
  kinases
• Akt is phosphorylated (active)
• Akt phosphorylation of FOXO sequesters it in
  inactive form
• Akt inhibits GSK-3, the general inhibitor of
  translation
• Inhibition of GSK-3 relieves translation
• Cells are prepared to proliferate

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MAP Kinase Signaling Pathways

• MAP kinases (mitogen-activated protein kinases)
  are protein-serine/threonine kinases
• Conserved across eukaryotic cells three groups
  of MAP kinases

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ERK Signaling Pathway

• ERK (extracellular signal-regulated kinase)
  family, first to be identified in MAPKs
• regulation of meiosis, mitosis, cell
  proliferation and differentiation
• Ligands growth factors, cytokines and viral
  infection, carcinogenic chemicals

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ERK Signaling Pathway

• ERK activation mediated by Ras, Raf, MEK kinase
  cascade
• Activation of Ras?activation of Raf protein
  serine/threonine kinase
• Raf phosphorylates and activates a second protein
  kinase called MEK (MAPK/ERK Kinase)
• MEK activates ERK? transcriptional activation

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ERK Signaling Pathway

• Ras guanine nucleotide-binding protein that
  function like a subunits of G proteins
• Ras is activated by guanine nucleotide exchange
  factors (GEF)
• Sosspecific GEF for Ras
• GTPase-activating proteines? GTP hydrolysis
• Ras-GDP becomes inactive

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ERK Signaling Pathway

• Grb2 SH2 domain containing protein associated
  with Sos
• RPTK activation by ligand recruits Grb2/Sos to
  membrane
• Grb2/Sos contacts Ras-GDP? GTP replaces GDP in
  Ras
• Ras-GTP activated and phosphorylates Raf
• Raf initiates a protein kinase cascade ?ERK
  activation

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ERK Signaling Pathway

• ERK goes to the nucleus, phosphorylates Elk-1
• transcriptional induction of immediate-early
  genes ( 100 genes)
• serum response element (SRE), recognized by
  transcription factors serum response factor (SRF)
  and Elk-1
• immediate-early genes encode transcription
  factors
• Activate downstream genes called secondary
  response genes
• Cell proliferation and growth

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ERK Signaling Pathway

• Specificity of MAP kinase signaling is maintained
  in part by their physical association on scaffold
  proteins
• For example, the KSR scaffold protein organizes
  ERK and its upstream activators Raf and MEK into
  a signaling cassette

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JAK/STAT Pathway

• Direct signaling from receptor to nucleus
• Ligand cytokines
• Receptors Janus Kinases (JAK), nonreceptor
  protein-tyrosine kinase
• STAT Signal Tansducer Activators of
  Transcription
• Transcription factors, contain SH2 domains that
  mediate binding to phosphotyrosine sequences
• STATs activated, dimerized, translocate to
  nucleus
• Activate transcription

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Notch Pathway

• direct cell-cell interactions during development
• Notch a receptor for signaling by transmembrane
  proteins (e.g., Delta) on adjacent cells
• Ligand binding ?proteolytic cleavage of cytosolic
  domain of Notch
• translocated into the nucleus
• converts a transcription factor (CSL in mammals)
  from a repressor to an activator
• Downstream genes code for other transcriptional
  factors
• Cell developmental differentiation

Minireview Notch signaling
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Integrins and Signal Transduction

• binding of integrins to the extracellular
• activation of FAK ( focal adhesion kinase), a
  nonreceptor protein-tyrosine kinase
• provides binding sites for Grb2-Sos complex,
  leading to activation of Ras/ERK, PI 3-kinase

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Integrins and Signal Transduction

• Rho subfamily of small GTP-binding proteins (Rho,
  Rac, and Cdc42) regulate organization of the
  actin cytoskeleton
• Rho family proteins promote actin polymerization