Marty Pagel, Ph'D' Department of Biomedical Engineering and Department of Chemistry

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BME 510 Biology for Biomedical
Engineering Lecture 32 and 33 Signalling Systems
Marty Pagel, Ph.D.Department of Biomedical
Engineering and Department of Chemistry Arizona
Cancer Center University of Arizona, Tucson,
AZ Email mpagel_at_u.arizona.edu Cell Phone
(520)-404-7049 Office Room 4949B, AZ Cancer
Center
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Typical External Signaling Methods
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General Format of Signaling
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How the Cell Responds
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Nuclear Receptors

• Often there is a link between a receptor and the
  promoter of a gene that is turned on or off.
• The receptor may be on the cell surface or in the
  nucleus.
• Intracellular receptors may respond to
  intracellular metabolites or to molecules that
  diffuse across the membrane.
• The nuclear receptor superfamily members respond
  to hydrophobic mediators, e.g. steroid hormones,
  thyroid hormones, retinoids, and vitamin D.
• Unlike water-soluble mediators, these are more
  stable in circulation and mediate longer
  time-frame regulatory pathways.

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Three Types of Cell-Surface Receptors

• Ion-channel linked receptors.
• G-protein-linked receptors.
• Enzyme-linked receptors.
• Many of these are or are linked to protein
  kinases.

Signal Transduction from the Cell Surface

• Signal transduction -- passing on information and
  applying it to specific functions.
• Two major direct signaling methods in cells.
• Protein kinases
• GTP-binding proteins

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The Two Major Pathways
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G-Protein-Linked Receptors
7 Transmembrane Segments Sometimes referred to
as the 7-Pass Transmembrane Receptors
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alpha or beta-gamma can activate other signaling
proteins.
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An Example Calcium release
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An Example of a Trimeric G-Protein Receptor
Olfactory Receptors

• Respond to 10,000 distinct smells.
• Receptors are G-protein-linked.
• Olfactorant binding activates adenylate cyclase.
• Increasing cAMP in the olfactory neurons opens
  cAMP-gated cation channels -- allows Na entry
  into the cell, which depolarizes the neuron and
  initiates a nerve impulse.

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Example Rod Photoreceptors (Rods)

• Rod cells mediate black and white vision,
  especially important at night.
• Light stimulates the hyperpolarization of the
  cell by stimulating cGMP phosphodiesterase,
  which closes cGMP-gated cation channels.
• The G-protein a subunit in this case is affected
  by conformational changes in the protein portion
  of rhodopsin (opsin).

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Rhodopsin
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Light-Activation to Close Cation Channels
When the cation channel is closed, internal Ca2
drops, which stimulates guanylyl cyclase to
regenerate cGMP levels.
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Desensitization
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Densitization of Rhodopsin Receptors
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Protein Kinase Family

• Two main types of protein kinases
  serine/threonine kinases and tyrosine kinases.
• 2 of all our genes are protein kinases.
• Signals can be amplified through cascades of
  kinases.

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Control by Phosphorylation
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Logic Functions
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Kinase Signaling Cascades
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Kinase Signaling Cascades
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Enzyme-Linked Receptors

• Receptor tyrosine kinases.
• Are tyrosine kinases themselves.
• Tyrosine-kinase-associated receptors.
• Receptorlike tyrosine phosphatases.
• Receptor serine/threonine kinases.
• Receptor guanylyl cyclases.
• Histidine-kinase-associated receptors.
• Transfers phosphate to one of its own histidine
  residues and then transfers the phosphate to
  another protein.

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Receptor Tyrosine Kinases

• Many are receptors for protein growth factors,
  including epidermal growth factor (EGF),
  platelet-derived growth factor (PDGF), fibroblast
  growth factors (FGF), hepatocyte growth factor
  (HGF), insulin, insulinlike growth factor-1
  (IGF-1), vascular endothelial growth factor
  (VEGF), macrophage-colony-stimulating factor
  (M-CSF), and nerve growth factor (NGF).
• They are also receptors for Ephrins (Eph
  Receptors).
• Ephrins may also be activated by the Eph receptor
  that it turns on.

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7 Subfamilies of Receptor Tyrosine Kinases
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Signal Transduction from Receptor Tyrosine Kinases

• Phospholipase C-? (PLC-?) --discussed below.
  Ultimately increases cytosolic Ca2.
• Kinases that in interact directly with the
  receptor tyrosine kinase.
• For example, Src tyrosine kinase, and
  phosphatidylinositol 3-kinase (PI3-kinase).
• These kinases have Src homology regions (SH2
  domains) that recognize specific phosphotyrosines
  on receptor tyrosine kinases.

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PhosphatidylInositol 3-kinase (PI 3-kinase)
Turns on proteins with PH domain
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Phospholipase C-? (PLC-?)

• Cleaves PI 4,5-bisphosphate, PI(4,5)P2 to IP3 and
  diacylglycerol.
• IP3 stimulates release of Ca2 from the ER.
• Diacylglycerol stimulates the Protein Kinase C
  (PKC).

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Protein Kinase C is a Calcium-Activated Kinase
PKC is a ser/thr kinase
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Other Serine-Threonine Kinases

• Protein kinase A (PKA) or cAMP-dependent kinase
• Ca2/Calmodulin kinases
• Calmodulin is a Ca2-dependent activator protein
  similar to troponin C -- It is a multipurpose
  Ca2 adapter.
• example MLCK (myosin light chain kinase)
• Mitogen-activated kinase (MAP-kinase)
• Protein kinase B (PKB)

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Cyclic AMP (cAMP)
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PKA (cyclicAMP-Dependent Kinase)
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Summary

• Signal transduction pathways carry information
  from the cell surface to inside the cell.
• Tyrosine kinases are often associated with growth
  factor receptors and couple into various signal
  pathways, including multiple kinases in a
  cascade.
• Other cell surface receptors use various enzymes
  to mediate the immediate response to stimulation
  of the receptor.
• Many of the enzymes coupled to cell surface
  receptors are Serine/Threnine kinases. These have
  multiple roles within the cell, including
  coupling Ca2 levels to specific responses within
  the cell.
• Phosphorylation by kinases and dephosphorylation
  applies binary logic to control many different
  functions of the cell.

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Jak-STAT Receptors

• Family of enzyme-linked receptors.
• Large, diverse class including receptors for
  cytokines hormones.
• Act through a group of tyrosine kinases called
  Janus kinases (Jaks).
• Regulate gene transcription through signal
  transducers and activators of transcription.
• In humans there are 4 Jaks and 7 different STATs
  identified.

Janus ancient roman god of beginnings (January,
Genesis) and endings.
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Cytokines

• In responding to cytokines and growth factors,
  the Jak receptors function largely in
  hematopoiesis (development of blood cells) and in
  antimicrobial immune responses.
• Cytokines are peptide hormones that include the
  interleukins, which orchestrate large scale
  changes in the character of an immune response,
  and interferons, which are oriented largely
  towards control of infection.

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Erythropoietin
Mouse Embryos, Day 13 Normal vs. No EpoR
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Multiple Signal Pathways Branch from the
Erythropoietin Receptor
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a-Interferon Jak-STAT Signaling
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a-Interferon Jak-STAT Signaling
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An Example Interferon
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Jak-STATs end a cells life in response to viral
infection

• Many viruses make double-stranded RNA (dsRNA) in
  the cells they infect.
• Mammalian cells can detect dsRNA and respond by
  producing interferon-a and interferon-ß.
• Activation of the Jak-STAT pathway induces a
  ribonuclease that destroys single stranded RNA
  and phosphorylates eIF-2.
• Protein synthesis in the cell is shut down --
  viral replication is shut down.

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Example Ras is a Related G-protein Receptor
Workshop Examples

• Ras is not trimeric, it is monomeric.
• Ras is equivalent to the a subunit.
• Ras activates the MAP-kinase pathway that
  stimulates cell growth.
• Mutation at Codon XII increases GTP binding,
  causing cancer

GTPase Activating Protein
Guanine Nucleotide Exchange Factor
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ExampleActivated a Subunit Activates Adenylate
Cyclase
Workshop Examples
Epinephrine receptors Glucagon receptors ACTH
Receptors
Continued on next slide
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Workshop Examples
ExampleActivated a Subunit Activates Adenylate
Cyclase
CREB cAMP-Response Element Binding Protein
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Mitogen Activated Kinase (MAP-kinase)
Workshop Examples
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Mitogen Activation Via Ras MAP Kinase
Workshop Examples
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5 Parallel Signaling Pathways
Workshop Examples