Intercellular and intracellular signals

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Intercellular and intracellular signals

• Primary signal molecules small, apolar mol's,
  unstable compounds, small and large water-soluble
  molecules hormons, paracrine-, autocrine- and
  juxtacrine (contact dependent) factors growth-,
  death-, survival and differentiation factors
• Receptorsintracellular and cell surface
  receptorsreceptors with DNA binding,
  transcription regulating activity, G-protein
  coupled receptors, ion channel-linked receptors,
  enzyme-linked receptors
• Second messengers (intracellular signal
  molecules)cAMP, IP3, DAG, Ca ions, protein
  kinases/phosphatases, ubiquitin-ligases,
  proteases, (scaffold proteins, cytoskeletal
  structures, motor proteins)
• TargetsDNA - general transcription factor
  complexes, histon proteins (nucleosome),
  metabolic enzymes, RNA processing mechanisms
  (splicesomes), extracellular matrix

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Growth-, survival- and differentiation factors

• On the basis of their biological activity we
  distinquish growth-, survival and differentiation
  factors
• Growth factors are mitogenic, elicit
  proliferation of the cell type,survival factors
  are needed for prolongation of life,differentiati
  on factors trigger irreversible changes in
  morphology and metabolism (some of them are
  death-factors, as terminal differentiation in
  certain cell types leads to programmed
  death)death-factors, triggering programmed
  death, lysis or necrosis of cells
• These factors are pleiotropic their effect
  depends on cell type, stage of differentiation,
  presence of other factors, etc.
• Survival factors can be growth factors for other
  cell types or cause apoptosis in another cell
  types.
• In general all these factors are referred to as
  growth factors (GF)

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Receptor types
Ion channel-linked receptors regulate the traffic
or transport of specific ions. Ligand binding
elicits opening of the channels. G-proteins
coupled receptors activate G-protein trimers and
indirectly (through the activity of G proteins)
induce the synthesis of second messengers, like
cAMP, IP3 and DAG. Ligand-binding induces
oligo-merization of enzyme-linked (and signaling
protein-linked) receptors, which leads to
activation of different enzymes and second
messengers.
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Receptor types
1. Receptor tyrosine kinases A number of growth
factor-, cytokine-, and some hormone receptors
exhibit tyrosine-kinase activity Ligand-binding
induces dimerization, activate the latent kinase,
which results in the mutual phosphorylation of
receptor subunits Second messenger molecules are
attracted to the phosphotyrosine residues of the
receptor The kinase also phosphorylates these,
activating these second messengers
platelet-derived growth factor
PDGF receptor
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Receptor types
1. Receptor tyrosine kinases A number of growth
factor-, cytokine-, and some hormone receptors
exhibit tyrosine-kinase activity Ligand-binding
induces dimerization, activate the latent kinase,
which results in the mutual phosphorylation of
receptor subunits Second messenger molecules are
attracted to the phosphotyrosine residues of the
receptor The kinase also phosphorylates these,
activating these second messengers In case of
these receptors the subunit(s) of the receptor
exhibit(s) kinase activity
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Receptor types
2. Tyrosine kinase-linked receptors Other
receptors are linked to kinases. The kinase is a
separate molecule, which associates with the
receptor after ligand-induced dimerization. The
kinases phosphorylate each other, subunits of the
receptor and second messenger molecules,
attracted to the active receptor
The receptor subunits have no kinase activity
receptor activation leads to kinase activation
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Channel-linked receptors

• Activation of these receptors leads to the
  opening of specific ion channels
• These receptors can be localized in the
  plasmamembrane or in intracellular membranes
  (eg. vitamin D3 receptor, IP3 receptor)
• ATP-gated receptors (P2X) playroles in
  contractility, tone, nociception, etc. are
  specific for calcium or sodium ions

P2X
vitamin D3 receptor
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Channel-linked receptors(ligand-gated channels)

• A number of neurotransmitters have receptors with
  ion channel activity (cys loop type of
  receptors). These can be specific for Cl ions
  (eg. GABA receptors, histamine receptor) or for
  cations (nicotinic acetylcholin R, 5HT).

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Channel-linked receptors(ligand-gated channels)

• NMDA (N-methyl-D-aspartic acid) receptor is one
  member of the glutamate receptor family. It has 4
  transmembrane domains. Splice variants are
  present in different cell types. It is a Ca(Na)/K
  pump.

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Mitogenic factors

• Each cell type have characteristic GF receptors.
  Only these can induce proliferation of the cell.
• Mitogenic factors are highly specific for cell
  type and degree of differentiation.
• The same factor might have other effects on other
  cell types (pleiotropic effect).
• (Tumor necrosis factor, TNF is a mitogenic factor
  for fibroblasts, but cytostatic on melanoma
  cells, induces differentiation of myeloid cell,
  dedifferentiation in chondrocytes, inhibits
  diffferentiation in myoblasts and kills different
  tumor cell types, oligodendrocytes, endothelial
  cells
• Mitogenic factors colony stimulating factors
  (CSF) for mieloid progenitors, interleukin-2
  (IL-2) T-cells, IL-6 B-cells, erythropoetin
  reticulocytes

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Signaling pathways
Receptors frequently induce several signaling
paths, which can interfere, synergize or modulate
each other. Some pathways are characteristic to
certain cell types. Membrane-associated
proteins are very important in
signaling. Mitogenic factors activate
themitogen-activated protein-kinase
(MAPK)pathways These are parallel
paths,serving different signals.All activate
transcriptionfactors (eg. Ap1)
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Signaling is an amplification cascade

• Each step amplifies the signal, more and more
  active molecules are generated
• Reversible phosphorylation is a key element of
  the signaling process
• Kinases phosphorylate tyrosine, serine,
  threonine amino-acids of proteins
• Activated proteins are either dephosphorylated
  by phosphatases or they are degraded (after
  ubiquinylation)

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Signaling of receptor tyrosine kinases

• Mitogenic (growth) factors activate one of the
  mitogen-activated protein kinase pathways, each
  representing a chain of proteins with kinase
  activity.

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Signaling of receptor tyrosine kinases

• The signal is amplified by each step. Activity of
  other enzymes are also modified by the kinases
  and other signal pathways might also be
  activated. The targets are DNA-binding
  transcription factors.

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Mutations of the receptors or signaling molecules
leads to disease

• Members of the FGF family of receptors from
  homo-and heterodimers with different
  specificities.
• Mutations affecting the ligand-binding domain
  might cause severe disease

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Signaling of kinase-linked receptors

• Some growth factor receptors signal through the
  activation of protein kinases.
• Ligand binding activates receptors. The kinase is
  a separate molecule (JAK), which associates with
  the activated receptor (ligand-induced
  dimerization). Adapter molecules (eg. STAT) are
  attracted to the phosphorylated receptor and get
  activated (phosphorylated) by the kinase.
• Activated adapters dimerizeand are translocated
  into the nucleus.
• Their DNA binding modifies activity of many
  genes (they function astranscription factors),

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Signaling of kinase-linked receptors
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Signaling of kinase-linked immune receptors

• Ligand-specific and signaling subunits build up
  these receptors. The same signaling subunit can
  serve many receptors with different ligand
  specificity.
• This may cause competition of different ligands
  for the receptors.

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Insulin receptor and signaling

• Activation of the insulin receptor triggers PI
  phosphorylation. An enzyme complex is assembled
  on the inner surface of the membrane.
• The activated kinases unlimately change the
  balance of glucose metabolism (in an opposite
  way what we learned aboutthe effect of glucagon)

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(No Transcript)
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Differentiation factors

• A differentiation factors like mitogenic
  factors highly specific for their target cells
• Receptors of differentiation factors is produced
  only in certain stages of differentiation and
  might disappear after one or several steps
• Differentiation factors are of different chemical
  nature mostly proteins, but steroids and
  retinoids are also potent morphogens
• A typical differentiation factor, transforming
  growth factor (TGF) is a member of a superfamily
  of proteins (activins, inhibins, TGFs, bone
  morphogeic proteins, BMPs, etc.).Their structure,
  signaling pathways, functions remained highly
  preserved during evolution

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TGF/TGF.R signaling

• Ligand binding activates the receptor
  (ligand-induced dimerization). Adapter molecules
  (SMADs) are attracted to the phosphorylated
  receptor and get activated (phosphorylated).
• TGF.R family members use specific Smad proteins
  for second messengers
• Smad1 to 3 and 5are signal trans-ducers, Smad4
  is a co-Smad, while Smad6 and-7 are inhibitors
  of Smad signals
• The pathway is verysimilar to the STAT pathway

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Survival factors and their receptors

• The population of certain cell types can be
  controlled at the level of proliferation or at
  the degree of survival
• 1. if GF is not present, the cells are living in
  G1 or G0 phase (no DNA synthesis). Proliferation
  is triggered by the presence of GF.
• 2. GF is always present, the cells are
  continuously proliferating, however, their
  lifespan is limited (terminal dfferentiation
  leads to apoptosis)
• Apoptosis of these cells can only be prevented
  by survival factors
• Hemopoetic stem cells (HSC) require stem cells
  factor (SCF),interleukin-3 (IL-3), and flt-3
  ligand (FL) for survival. Absence of these
  factors leads to death, not only growth-arrest.
• Survival factors NGF (nerve GF) neurons, IGF-1
  (insulin-like GF) many cell types, endothelin
  endothel cells, BAFF B-cells, leptin eozinophil
  granulocytes, neurotrophins neurons, embryonic
  stem cells, VEGF (vascular endothelial growth
  factors) capillary endothel, retina endothel
• Survival factor receptors belong to different
  receptor families (mostly GF.Rs)

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Death receptors
Certain cell types (eg. autoreaktív T and B
cells) must be killedActivation of death
receptors (fas, TNF.RI, NGF.R, TGF.R, etc.) leads
to programmed cell death in a cell
type-specific way. The same receptors might
function as growth factor receptors in other cell
types
death pathway survival pathway
proliferation
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Wnt signal a special pathway

• Wnt signal also serves as a differeniation signal
  (ventral-dorsal) during ontogenesis
• The receptor is a 7TM domain GPCR, which
  activates more than one pathway
• In the absence of the ligand beta catenin is
  degraded. Activation of the receptor leadsto
  stabilization of catenin, allowing
  nucleartransport and gene regulation by
  activatingtranscription factor(s).

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Catenin is a multifunctional protein
beta catenin is associated with cytoskeletal
structures linking cells of the same cell type
together
The Axin-APC-GSK complex (inducing
phosphorylation-linked ubiquitination) degrades
only free beta-catenin molecules, not interfering
with the structural function of the protein
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Wnt signal and differentiation

• Wnt can not activate its receptor in the
  posterior segment of the embryobecause of
  interference of other factors

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The balance of kinases and phosphatases

• Signaling is influenced by the balance of kinases
  and phosphatases. Rafts (membrane microdomains)
  can influence this balance while receptor and
  kinase are raft-associated, the phosphatase is not

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The balance of kinases and phosphatases

• Signaling is influenced by the balance of kinases
  and phosphatases. This balance is cell type
  specific and can inhibit the generation of the
  signal
• Though the ligand inter-acts with its cognate
  receptor no signal is generated
• SHP-1 enzyme de-phosphorylates the subunits of
  the receptor and the second messenger
  molecules

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Scaffold proteins
Scaffold proteins are matchmakers,they catalyze
the interaction of signal molecules Molecules
participating in subsequent steps of the signal
are bound to the surface of scaffold proteins,
recognizing and modifying each other Heat shock
proteins do the oppositetry to inhibit
interaction of signaling molecules in the absence
of ligand, preventing false signals
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Pathways of TNF.R signaling
Members of the TNF.R super-family can signal
death as well as survival, proliferation,
activation and differentiation. In different
cell types different pathways dominate, other
might be missing The pathways are under
elaborate control mechanisms.
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Differentiation steps

• In a cell at a certain stage of its
  differentiation
• - a set of transcription factors regulate the
  activity of genes,
• - a set of its genes show characteristic pattern
  of histone and DNA modification
• - the hnRNAs are spliced with characteristic
  splicing factors, resulting in a set of splice
  variant mRNAs,
• - the proteins are modified by characteristic
  processing enzymes
• - it results in a characterisitc enzyme activity
  pattern, metabolic activity and morphology
• - life span of the proteins is regulated by
  characteristic factors
• - the cell secretes specific proteins and
  exhibits a characterisitc set of receptors.
• The metabolic ativity and the morphology allows
  the cell to perform specific tasks, determines
  the way how to communicate with other cells and
  how to respond to external signals.
• Depending on the signals detected by its
  receptors, the cell can have different fates
  proliferation, differentiation, activation or
  death

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Differentiation steps

• The differentiation factor interacting with the
  receptor of the cells induces a signal process
  resulting in specific changes
• - in the activity of enzymes (different
  phosphorylation patterns, different activities),
• - in the composition of transcription factors,
• - in the modification of histones and DNA,
• - inducing synthesis of a new sets of hnRNAs,
• - splicing of these hnRNAs with a new set of
  splicing factors,
• - resulting in the synthesis of new splice
  variant proteins and new sets of enzymes,
  receptors, ECM proteins,
• - changing the pattern of protein-modifying and
  protein-degrading enzymes,
• - resulting in different metabolism,.
• The new metabolic activity might mean changed
  morphology and new functions. The cell starts to
  produce new receptors and signal molecules,
  different ECM its communication with other cells
  has also changed.
• In many case the differentiation process is
  irreversible.

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Signal and co-signal
In cell-fate decisions signals frequently need a
corroborating co-signal to be effective In other
cases, the presence of a co-signal alters the
meaning of the original signal In immune cells
antigen co-signal survival, activation,
proliferation antigen alone apoptosis or
anergy
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Juxtacrine signal and ECM signal
In cell differentiation ligands originating from
neighboring cells (cell adhesion molecules,
juxtacrine factors) or the extracellular matrix
(ECM) play very important roles
The signal is usually bi-directional. Both cells
differentiate (in the same or different
directions)
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ECM-triggered signaling
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ECM-triggered signaling

• The cells produce cell-type specific ECM
  molecules. These proteins activate different
  signaling pathways in the signaling and the
  neighboring cells. These signals harmonize cells
  of a certain tissue, create links between
  different cell types or induce differentiation of
  cells.
• Similar cells are linkedtogether by
  homofil-,different cell types by heterofil cell
  adhesionmolecules (CAMs)

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Cross-talk of signaling pathways

• Cells are exposed to simultaneous signals. The
  signaling pathways and signaling molecules
  modulate each other they synergize, interfere or
  modify each other.
• The effect of a certain factor depends on the
  presence of all other factors

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The EGF.R signaling pathways
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Alternative pathways result in different effects
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Survival factors
Apoptosis-prone cells survive only in the
presence of survival factors Neurons NGF,
neurotrophic factors, hemopoetic SC SCF, IL-3,
flt3-L The survivalfactors block
thepro-apoptoticsignal cascades,protecting the
life of the cells