1 Signal transduction

• Definiton: Oncogenes are parts of a cell's genome that, when excessively activated, promote the transition from normal cell growth behavior to unrestrained tumor growth

• Cause stop of proliferation in wild type cells ( if on contact)

• Proto-oncogenes are genes that normally help cells grow and divide to make new cells, or to help cells stay alive. When a proto-oncogene mutates (changes) or there are too many copies of it, it can become turned on (activated) when it is not supposed to be, at which point it's now called an oncogene. When this happens, the cell can start to grow out of control, which might lead to cancer.

  A proto-oncogene normally functions in a way much like the gas pedal on a car. It helps the cell grow and divide. An oncogene is like a gas pedal that is stuck down, which causes the cell to divide out of control.

• Viral oncogenes transform normal cells by retro viruses into cancer cells

  the viruses contain small pieces of human DNA

  can be caused by shortenings or point mutations

Influences transformation rate

The massive overexpression of an oncogenes can cause a transformation of cells.

The combinatorial expression of several oncogenes often has a synergistic (working together) effect.

Often oncogenes controlling proliferation and affecting cell fate work together.

Proto-oncogenes encode factors of signal transduction pathways

Signal transduction controls almost all cellular responses => key mechanism

Signal transduction can induce:

• determination of tissue/expression domains

• differentiation of different tissue types

• selection of individual cells from

  precursor tissues

• Proliferation control

• Targeted cell death

• Cell migration

• Altered metabolism

Induction/determination depends on the competence of the target cell

• All mechanisms of signal transmission depend on the

  competence of the target cell

• Thus, the same signal may have different "responses“ in

  different cells

• A gene can be activated by different signals. Some use the same pathways, others use overlapping pathways

• Due to different additional factors (co-factors), the expression of the same gene has different consequences

• Examples: notch signaling (lateral inhibition, lineage, boundary), dpp signaling (gradient: target cells have different competencies -> react different)

Slow and rapid cellular responses to an extracellular signal

Signal peak followed by revision (fast turn on/off)

Examples

• Protein phosphorylation by a kinase followed by dephosphorylation by a phosphatase

• Activation by binding of a co-factor: G-proteins with an exchange GDP (inactive) into GTP (active).

• Protein complex formation: association at the plasma membrane

• Protein activation by protein cleavage (Active for quite some time)

• Change of subcellular localization: movement into nucleus (fast and easy reversible)

• Release of second messengers: release of Ca2+ from the ER

• Destruction of protein of the signal transduction pathway

Signal integration and/or positive feedback loops cause an cellular response

• Y makes sure that it doesn’t react on accident/ wrong signal -> needs combination of two signals (strong signal)

• Sigmoids graphic

• Positive feedback loops can induce an all or nothing response

• Positive feedback can translate a transient signal into a persistent change

• An allosteric binding requirement can result in a sigmoidal activation thereby inducing a threshold (example: parallel binding of 4 proteins required)

• can cause a requirement for strong stimuli

• Make sure that signal is turned down after the peak

• Negative feedback loops counteracts the effect of the stimulus (To create requirement for higher dose or to show an adaptation to a signal: the effect needs a higher dose for each additional activation)

Ion-channel coupled receptors

• acetylcholine

• Heart pacemaker cell

• Salivary gland cell (-> secretion)

• Skeletal muscle cell (-> contraction)

G-Protein coupled receptors

• G-Protein coupled receptors (GPCRs) form the largest group of receptors

  • More than 800 in human genome

• GPCRs are involved hormone, neurotransmitters, smell, taste and sight

• All GPCRs have a similar structure: many (often seven) transmembrane domains

  • An extracellular ligand binding domain

  • An cytoplasmic interaction domain for the G-protein interaction

• Trimeric G-Protein coupled receptors

• All GPCRs bind to a trimeric G-protein

• Trimeric G-proteins bind specifically a group of GPCRs

• The trimeric G-protein couple the GPCR with a channel or enzymes

• Activation after binding to the GPCR is based on the exchange of GDP with GTP -> exchange causes dramatic resolving and separation of subunits.

  • Both the a subunit and the b y subunit activate independent downstream signaling ways

  • The a subunit hydrolyzes GTP to GDP by itself, but this inactivation can be fastened by the interaction with a regulator of G protein signaling (RGS) the functions as a GAP (GTPase activating enzyme)

• Some G-proteins regulate the production of

  cyclic AMP (cAMP)

• The synthesis of cAMP is regulated by adenylyl

  cyclase

• cAMP is essential for a diverse set of cellular responses

• Adenylyl cyclase is also multitransmembrane protein in the plasmamembrane

• cAMP functions as a diffusible co-factor of the cyclic AMP dependent protein kinase A (PKA)

• cAMP is inactivated by the phosphodiesterases, thereby inhibiting the activation of PKA in the cell

• cAMP effects mainly via protein kinase A (PKA)

• The inhibitory subunit of PKA restricts the the localization of PKA

• cAMP bind to the inhibitory subunit of PKA

• The cAMP binding allows the release of the catalytic active kinase subunit

Activation of PKA = allosteric activation

• Activated a subunit of G-protein activates adenylyl cyclase

• cAMP bind to inhibitory subunit of PKA

• Activated PKA is released: It phosphorylates many cytoplasmic proteins -> cytoplasmatic response

  • Among those are phosphodieseterases that function as a negative feedback loop by inactivating cAMP

• Activated PKA moves in some cells also into the nucleus

• It activates the cyclic AMP response element binding protein (CREB) by phosphorylation

• Resulting in altered gene expression

• Some activated G-protein can stimulate the activity of phospholipase C (PLC)

• PLC release the head group (IP3) of certain membrane lipids and the remaining Di-acylglycerol (DAG) in the membrane

  => both are second messenger signaling molecules

• IP3 stimulates the release of Ca2+ ions from the ER which functions as a co-factor of Protein kinase C (PKC)

• DAG activates Protein kinase C (PKC)

IP3 stimulates the release of Ca2+ ions from the ER that can move fast within the cell

• Ca2+ bind to calmodulin

• Ca2+ calmodulin bind to the inactive CaM kinase

• The autophosphorylation of CaM results in the fully active version of CaM

• Release of Ca2+ calmodulin and de- phosphorylation inactivates CaM

• Ca2+ ions bound to calmodulin bind to Ca2+ pumps that pump the Ca2+ ion backwards into the ER with a delay (negative feedback loop)

Receptor tyrosine kinases (RTK)

• growth, proliferation, development…

• kinases phosphorylate amino acids: serine, threonine and tyrosine

RTKs activate independent signaling pathways

Ligand binding cause dimerization and auto-phosphorylation of RTK

Adaptor proteins bind to tyrosine phosphorylated receptors

RTK Signaling causes many effects (examples: proliferation, growth, survival, metabolism, differentiation)

GTP bound Ras activates Raf kinase starting a phosphorylation cascade

• MAPK/ERK stimulates gene expression via Ets and SRF transcription factors

  • Erk phosphorylates Ets

  • Erk phosphorylatesRSK

  • RSK phosphorylates SRF (serum response factor)

  • Ets activates together with SRF gene expression

• Many regulated enhancers contain ETS and SRF Binding sites

Loss of EGF Signaling cause apoptosis in Drosophila

• RTK Torso is ubiquitous expressed

• The ligand Trunk is only present in perivitelline space at the anterior and posterior end of the embryo

• So Torso is only activated at both poles

Signaling through RTKs activates PKC

Signaling through RTKs activates PI3 kinase

PI3,4,5 triphosphate (PI(3,4,5)P3) membrane lipids enable Akt activation

• Activated PI 3 kinase synthesizes PI3,4,5 triphosphate (PI(3,4,5)P3) membrane lipids

• PI(3,4,5)P3 functions as a binding site of protein with a pleckstrin homology domain (PH)

• PH domain proteins protein kinase B (Akt) and phosphoinositide-dependent kinase (PDK1) bind

• Resulting in the activation of Akt by phosphorylation

• Akt has many targets among them proteins controlling cell death (apoptosis)

example of signalling cascade (exception)

• Ephrin receptors autophosphorylate after ligand binding

• Phosphorylated tyr of the receptor enable Rho GEF interaction

• Rho GEF catalyzes the GDP to GTP exchange of Rho, which becomes active

• Local activation of Rho cause a breakdown of the local cytoskeleton, resulting in the repulsion of filopodia of growing axons

• beside Rho, also Rac and Cdc42 function as regulatory proteins of the cystoskeleton

Fibroblast growth factor

• 3 signal transduction pathways are stimulated: Ras/Raf/MAPK path

• Activation induces activity of various transcription factors

• Gene expression of target genes

• Cellular responses: Morphology, survival, proliferation

FGF example (not detail)

• Filopodium (highly dynamic) with FGF receptor

• activation of Btl in the tip cell(s) leads to the formation of filopodia and laminopodia in these cells

• The activated cells stretch themselves and with them the following cells of the stalk

• Btl activation in the tip cell induces activation of Delta (Notch ligand)

  The binding of the Delta ligand to its Notch receptor in stalk cells blocks change in fate.

  The branching outgrowth stretches further by cellular intercalation, the cells change their position relative to each other.