Lecture 9 - Tyrosine Kinase Signaling Flashcards
What are the 2 types of tyrosine kinases?
- Receptors
2. Receptor-associated
List 5 receptor tyrosine kinases.
- Insulin receptor
- IGF-1 receptor
- Nerve growth factor receptor
- Epidermal growth factor receptor
- Platelet derived growth factor
+ many more growth factor receptors
What must be true of receptor associated tyrosine kinases?
They are soluble in the cytoplasm
What do the receptor tyrosine kinases have in common? 3 parts
- 1 single transmembrane domain
- Elaborate, large, and globular extracellular domain
- Intracellular tyrosine kinase catalytic domain
Where is the catalytic domain of receptor tyrosine kinases located?
Proximal to the membrane on the cytosolic side
Describe the 4 steps of receptor tyrosine kinase activation.
- Ligand binding induces receptor dimerization through various mechanisms
- Dimerization activates the tyrosine kinase domain of the receptor by moving the regulatory binding from the active site
- Tyrosine kinase INTERmolecularly trans-autophosphorylates the receptor mostly OUTSIDE of the catalytic domain
- Substrate phosphorylation
Which ONLY receptor tyrosine kinase is a dimer? How?
What is also unique about this receptor’s functioning?
Insulin receptor is covalently dimerized
Also phosphorylates insulin receptor substrates (IRS) on tyrosines, creating even more phosphotyrosine binding sites, on top of the usual binding sites on the receptor itself
Is the trans-auto-phosphorylation of receptor tyrosine kinase necessary for proper substrate phosphorylation?
YUP
What is the most fundamental difference between tyrosine kinases signaling strategies and Ser/Thr kinases signaling strategies?
- Ser/Thr kinases: presence of P on substrates results in a conformational change which activates or inactivates(couple of exceptions)
- Tyr kinases: some small conformational changes, but mainly the cytosolic phosphotyrosines serve as binding sites for phosphotyrosine binding proteins
What are 3 examples of phosphotyrosine binding proteins?
- PI3-K using a p85 adaptor
- GAP = GTPase Activating Protein
- PLPC-gamma
Describe the specificity of the phosphotyrosine binding sites and give a supporting argument.
Each binding protein has a VERY specific recognition 5 AA sequence including the phosphotyrosine
Synthetic peptides, as short as 5 AAs, containing a P-tyrosine can block binding to SPECIFIC binding sites on the receptor
Why did researchers originally think the tyrosine kinases had low affinity and specificity? How is this overcome by the tyrosine kinase?
Because they have low affinity and specificity at their catalytic site, not their phosphotyrosine binding sites:
- Deal with lack of specificity by having the high specificity at the 5 AA sequences
- Deal with lack of affinity by bringing in the substrates to the enzyme, creating an artificially high substrate concentration to work with the high Km of the catalytic site
What parts of the phosphotyrosine binding proteins bind to the binding sites?
Src homology 2 domains (SH2)
2 types of receptor associated protein tyrosine kinases?
- Src
2. Jak/STAT
To what kind of receptors do receptor associated protein tyrosine kinases bind? 6 examples?
Cytokine receptors
Eg: GH receptor, prolactin receptor, interferon receptor, erythropoietin receptor, G-CSF receptor, IL receptors
To what do the receptor associated protein tyrosine kinases bind on the receptors?
Tyrosine kinase binding domain on the cytosolic side
Do receptors that receptor associated protein tyrosine kinases bind to also form dimers?
YUP
Describe the 4 steps of receptor associated tyrosine kinase activation. (2 options for the first 2 steps)
- Ligand binding induces receptor dimerization through various mechanisms
- Dimerization recruits a tyrosine kinase binding to the receptor
OR - Tyrosine kinases already bound to each receptor
- Ligand binding induces receptor dimerization
- Tyrosine kinases INTERmolecularly trans-autophosphorylate each other and then the receptor mostly OUTSIDE of the catalytic domain
- Substrate phosphorylation (same process as receptor tyrosine kinases)
What is another name for the JAK kinases?
Janus kinases
How many kinase catalytic domains do Janus kinases have? What to note though?
2 (only 1 is particularly active though)
Do the receptor associated tyrosine kinases have an SH2 domain? What to note though?
Yes, but that is not what they use to bind the receptor (use dimerization type domains instead)
What are the 4 subtypes of JAK kinases?
- Tyk2
- Jak1
- Jak2
- Jak3
What is particular about Jak3?
Tissue specific: expressed in myeloid and lymphoid tissues
What does substrate specificity for receptor associated tyrosine kinases depend on?
The nature of the 2 tyrosine kinases in the dimer
What is the main difference between the activity of the receptor tyrosine kinases and the receptor associated tyrosine kinases?
The receptor associated ones first trans-auto-phosphorylate each other before trans-auto-phosphorylating the receptor
What are STAT proteins?
Signal Transducers and Activators of Transcription
How do STAT proteins work? 4 steps
- SH2 domain containing proteins and are recruited to phosphotyrosines binding sites created by Jaks or srcs
- Tyrosine kinases phosphorylate the STAT proteins
- STATs dissociate from the receptor and dimerize via their SH2 domains binding to the phosphotyrosine on the other STAT
- STAT dimer migrates to the nucleus and other gene regulatory proteins bind to it
What are srcs? How do they work?
Same role and functioning as Jak
SH2 domain containing tyrosine kinases that are implicated in many cancers
Can Jak and src dimerize together?
YUP
Can srcs also recruit STATs?
YUP
What is the built in OFF mechanism of the src and JAK signaling pathways? What is this called?
Negative feedback: STAT activates SOCS = Suppressors of Cytokine Signaling = will block further activation at the receptor by either binding at the SH2 binding sites or interfering with phosphorylation
What is a built in OFF mechanism of all tyrosine kinases? 2 types? Eg?
Tyrosine phosphatases
2 Types:
- Bound to the receptor (eg: binding site on platelet derived growth factor receptor for an oncogene called Syp = tyrosine phosphatase)
- Soluble phosphatase with SH2 binding site to remove phosphates from either the receptor itself, on substrates of tyrosine kinases still bound to the receptor, or substrates of tyrosine kinases that are soluble in the cytoplasm
Describe the PI 3 K cascade once activated.
PI3K phosphorylates PIP2 => PIP3 => activated PDK 1 => activates PKB
Other than tyrosine kinase, what else can activated PI 3 kinase?
Ras
How does STATs dissociating from the receptor and dimerizing via their SH2 domains binding to the phosphotyrosine on the other STAT allows them to go the nucleus?
Dimerization reveals its nuclear localization signals
What does trans-autophosphorylation mean?
The phosphorylation by a protein of a residue on an identical protein
