Introduction to Signaling
Networks
Biophysics 6702, February 2013
Jonathan P Butchar
jon.butchar@osumc.edu
Outline
• General Signaling Concepts
• Types of Signaling
• Signaling Components
– Receptors and Ligands
– Downstream Signaling
• Adaptor and Effector molecules
• Example Signaling Network: FcγR
Outline
• General Signaling Concepts
Different signals and signal combinations
lead to different outcomes
Figure 15-8 Molecular Biology of the Cell (© Garland Science 2008)
A single signal can lead to different outcomes
Figure 15-9 Molecular Biology of the Cell (© Garland Science 2008)
How?
Figure 15-9 Molecular Biology of the Cell (© Garland Science 2008)
Strength of signal can determine outcome
Figure 15-10 Molecular Biology of the Cell (© Garland Science 2008)
How else could a single signal lead to different
outcomes?
Figure 15-10 Molecular Biology of the Cell (© Garland Science 2008)
Slow and fast response times
Figure 15-6 Molecular Biology of the Cell (© Garland Science 2008)
Different signals can converge on one target
Figure 15-20 Molecular Biology of the Cell (© Garland Science 2008)
Crosstalk between signaling pathways
Figure 15-66 Molecular Biology of the Cell (© Garland Science 2008)
Cells can adapt to signaling
Figure 15-51 Molecular Biology of the Cell (© Garland Science 2008)
How else might cells adapt?
Figure 15-51 Molecular Biology of the Cell (© Garland Science 2008)
Signals can be amplified
Figure 15-28a Molecular Biology of the Cell (© Garland Science 2008)
or dampened
Figure 15-28c Molecular Biology of the Cell (© Garland Science 2008)
Primary and secondary responses
Figure 15-15 Molecular Biology of the Cell (© Garland Science 2008)
Primary and secondary responses
How to tell the difference?
Figure 15-15 Molecular Biology of the Cell (© Garland Science 2008)
Outline
• Types of Signaling
Figure 15-4a Molecular Biology of the Cell (© Garland Science 2008)
Don’t forget Autocrine signaling
Figure 15-4b Molecular Biology of the Cell (© Garland Science 2008)
Figure 15-4d Molecular Biology of the Cell (© Garland Science 2008)
Figure 15-4c Molecular Biology of the Cell (© Garland Science 2008)
Direct Transmission
Figure 15-7 Molecular Biology of the Cell (© Garland Science 2008)
Outline
• Signaling Components
– Receptors and Ligands
Figure 15-3a Molecular Biology of the Cell (© Garland Science 2008)
Figure 15-3b Molecular Biology of the Cell (© Garland Science 2008)
Numerous types of ligands
•
•
•
•
•
•
•
Peptides / Proteins
Steroids
Nucleotides
Fatty Acids
Gases
Mechanical Forces
temperature, etc
How do you get from ligand binding to an
intracellular response?
• Ion fluxes
• G-protein activation
• Enzyme activation (e.g., Phosphorylation)
Na+, K+, Ca2+, ClThere are voltagegated ion channels
too
e.g., Cystic Fibrosis is caused by defects in a Chloride channel
Figure 15-16a Molecular Biology of the Cell (© Garland Science 2008)
G-proteins: Guanine nucleotide-binding proteins
Figure 15-16b Molecular Biology of the Cell (© Garland Science 2008)
Enzyme-containing or enzyme-linked
Figure 15-16c Molecular Biology of the Cell (© Garland Science 2008)
Example: Receptor Tyrosine Kinases
Figure 15-53a Molecular Biology of the Cell (© Garland Science 2008)
Inhibition of Receptor Tyrosine Kinases
Figure 15-53b Molecular Biology of the Cell (© Garland Science 2008)
Inhibition of Receptor Tyrosine Kinases
How else could you do this?
Figure 15-53b Molecular Biology of the Cell (© Garland Science 2008)
Receptors can be locked in an inactive state
Figure 15-14b Molecular Biology of the Cell (© Garland Science 2008)
…and then unlocked by a ligand
Figure 15-14c Molecular Biology of the Cell (© Garland Science 2008)
Outline
• Downstream Signaling Components
– Signaling, Adaptor and Effector molecules
Signaling molecules transduce receptor activation
Figure 15-54 Molecular Biology of the Cell (© Garland Science 2008)
Common domains of signaling molecules
Proline-rich
Figure 15-22 Molecular Biology of the Cell (© Garland Science 2008)
Did I mention this is a really good book?
What molecules transmit these signals?
• Kinases and Phosphatases
• GTP / GDP
Kinases and phosphatases
e.g., Receptor Tyrosine Kinase:
a receptor AND a kinase
Figure 15-18a Molecular Biology of the Cell (© Garland Science 2008)
Guanosine TriPhosphate and
Guanosine DiPhosphate
Remember the G-protein coupled receptor
Figure 15-18b Molecular Biology of the Cell (© Garland Science 2008)
Trimeric G-proteins
• 3 subunits
– α, β, γ
Figure 15-32 Molecular Biology of the Cell (© Garland Science 2008)
Monomeric G-proteins
• Most well-known is Ras
– small GTPase
– downstream Raf binds
only GTP-Ras, which
phosphorylates and hence
activates Raf
GTPase-activating
protein
Figure 15-19 Molecular Biology of the Cell (© Garland Science 2008)
Guanine nucleotide
exchange factor
How do all these things get together?
Figure 15-21c Molecular Biology of the Cell (© Garland Science 2008)
$125.99 at Amazon.com.
Scaffolding proteins can aid the
interaction of signaling molecules
Figure 15-21a Molecular Biology of the Cell (© Garland Science 2008)
Binding occurs through characteristic domains
• PTB binds
phosphotyrosine
• SH2 binds
phosphotyrosine
• SH3 binds prolinerich domains
• PH binds
phosphoinositides
Figure 15-22 Molecular Biology of the Cell (© Garland Science 2008)
Proline-rich
Signaling molecules can also associate
directly with receptors
Figure 15-21b Molecular Biology of the Cell (© Garland Science 2008)
Example: from G-proteins
to gene transcription
•
•
•
•
Activated receptor
Activated G-protein
Activated Protein Kinase A
The Effector, an activated
transcriptional modulator
Figure 15-36 Molecular Biology of the Cell (© Garland Science 2008)
Lines are blurry at times
• Some membrane-bound receptors (e.g., glucocorticoid
receptors) can go to the nucleus and regulate gene
transcription
– Both a receptor and an effector
• Phosphorylation can sometimes deactivate rather than
activate a protein (e.g., the NF-κB inhibitor IκBα)
Outline
• Example Signaling Network: FcγR
Fcγ Receptors bind the Fc portion of IgG
FcγRIIb
(CD32b)
FcγRIIa
(CD32a)
FcγRI
(CD64)
FcγRIIIa
(CD16)
γ γ
γ γ
Inhibitory
ITAM
ITIM
Activating
ITAM
ITAM
ITAM
ITAM
membrane
Fcγ Receptors bind the Fc portion of IgG
• Rituximab, Herceptin, etc
• Autoantibodies (e.g., rheumatoid arthritis)
Phagocytosis
What kind of signaling is this?
http://www.whfreeman.com/immunology/CH01/figure01-04a.gif
Immune
Complex
Y
FcγR
Sos
• FcγR must be clustered
• Phosphorylation drives
downstream events
Y
FcγR activation
Grb2 Shc
-P
– ImmunoTyrosine-based
Activation Motif
Syk
• ITIM phosphorylation
dampens FcγR activity
Btk
Ras /MAPK
-P
Ca2+ flux
Src
PI3-K
PtdIns 3,4,5P
3
Vav
Akt
NF-κB
Phagocytosis
Gene Transcription
Y
Immune
Complex
Y
Downstream signaling: Ras and PI3K
Sos
FcγR
Grb2 Shc
-P
-P
Src
PI3-K
Syk
Btk
Ras /MAPK
Ca2+ flux
PtdIns 3,4,5P
3
Vav
Akt
NF-κB
Phagocytosis
Gene Transcription
Ras signaling review in under 10 seconds…
• G-protein
• small GTPase
Figure 15-60 Molecular Biology of the Cell (© Garland Science 2008)
2002 edition searchable for free at
http://www.ncbi.nlm.nih.gov/books/NBK21054/
Quick PI3K signaling review
(phosphoinositide 3-kinase)
Requires membrane localization
Figure 15-64 Molecular Biology of the Cell (© Garland Science 2008)
Y
Immune
Complex
Y
There’s crosstalk between Ras and PI3K
Sos
FcγR
Grb2 Shc
-P
-P
Src
PI3-K
Syk
Btk
Ras /MAPK
Ca2+ flux
PtdIns 3,4,5P
3
Vav
Akt
NF-κB
Phagocytosis
Gene Transcription
Ras
src
P
P
P
PIP2
PIP3
P
5
5
Shc
Sos
Grb2
4
PI-3K
Syk
PLC
Erk
4
3
Ca++
Vav
Rac
Btk
Actin
Polymerization
Phagocytosis / ROS / Cytokine
Akt
Y
Immune
Complex
Y
How could we modulate FcγR activity?
Inflammatory Cytokines
(IL-1, TNFα etc)
Sos
FcγR
O2-
Shc
Grb2
-P
-P
Src
Syk
Btk
PI3-K
PtdIns3,4,5P3
Ras /MAPK
Ca2+ flux
Vav
Akt
Phagocytosis
Gene Transcription
Remember, activating and inhibitory receptors
FcγRIIb
(CD32b)
FcγRIIa
(CD32a)
FcγRI
(CD64)
FcγRIIIa
(CD16)
γ γ
γ γ
Inhibitory
ITAM
ITIM
Activating
ITAM
ITAM
ITAM
ITAM
membrane
Summary
• Cells and groups of cells possess mechanisms to
generate and respond to signals
• Signaling can be autocrine, paracrine, endocrine,
synaptic, electrical or mechanical
• Receptors sense numerous types of stimuli and
begin cascades that lead to cellular responses
• Observed responses represent an integration of
stimuli, both past and present