L.2. Mario. PTMs, growth factor receptors and kinase activity.

Ubiquitination.

Tagging proteins with smaller proteins called ubiquitous, marking them for degradation by the proteasome.

Some proteins are not fully degraded, but form functional cleaved products.

Can regulate activity of some inflammatory pathways.

Protein phosphorylation.

One of the main mechanisms through which signals are transduced.

Phosphate is a fundamental chemical component in signalling.

Phosphorylation = acquisition of a phosphate group by a protein. Post-translational protein modification.

Facilitates protein-protein interactions: initiates a chain reaction.

• The discovery of phosphorylation led to the Nobel prize in physiology and medicine (1992).

Signal transduction is where: each relay molecule in the signal transduction pathway changes the next molecule in the pathway.

Dephosphorylation.

Protein phosphatases act as critical ‘off switches’ in signal transduction by removing phosphate groups from their target proteins.

Kinases are

The opposite of phosphatases in function, they add a phosphate group to a molecule.

Some phosphatases are particular to certain kinases: DUSPs (Dual-specificity phosphatases) dephospharylate

MAPKs (mitogen-activated protein kinases)

by removing phosphate groups from both threonine and tyrosine residues.

Some phosphatases are particular to certain kinases: PTPN2 - (Protein Tyrosine Phosphatases, Non-Receptor type) dephosphorylate…

STATs (Signal Transducers and Activators of Transcription) proteins and their upstream Janus kinases (JAKs).

Receptor tyrosine kinases (RTKs)

• The human kinome has around 90 tyrosine kinases: enzymes that phosphorylate other proteins. Of these, around 60 are receptors. Receptors with intrinsic kinase activity.

• Structure comprised of: An extracellular ligand binding domain, a single transmembrane helix, and an intracellular region that contains a tyrosine kinase domain (TKD).

How RTK phosphorylates amino acid residues.

• When a ligand binds, two RTK molecules come together, dimerising.

• Their receptor tails have intracellular kinase domains that then phosphorylate tyrosine (tyr) amino acids on the other receptor.

Ligand — RTK dimerisation — Receptor tails intrinsic kinase activity activated — Tyr phosphorylation.

Phosphorylated tyrosines on the RTK become docking sites for signalling proteins.

Different proteins recognise different phosphorylated tyrosines using domains such as:

• SH2 domains: bind phosphotyrosine (pTyr)

• PTB domains: also recognise phosphorylated tyrosine containing sequences.

EGF-R family is a family of RTKs. 4 main members?

HER1-4 or ErbB1-4.

EGF-Rs and cancer.

• EGF-R mutations affecting ATP-binding pocket result in hyperactivation of the kinase and downstream signalling, oncogenic.

• Overexpression of the HER2 gene occurs in approximately 15-30% of breast cancers.

• Increased local concentration of the receptor = increased RTK signalling overwhelms physiologic regulation.

• EGF-R: blockade with monoclonal antibodies used in cancer therapy.

Erythropoietin (EPO) function.

Erythropoietin (EPO) is a hormone that stimulates proliferation of progenitor cells in bone marrow. It increases the number of erythrocytes (red blood cells).

Kidney releases EPO, increasing erythrocytes to increase oxygen carrying capacity. Blood haemoglobin levels increase.

Body, kidney is adaptive, kidney increases the release of erythropoietin when blood oxygen levels are low (hypoxia).

EPO-R

This is a receptor that activates tyrosine kinases but does not possess intrinsic RTK activity.

EPO-R is a cytokine receptor associated with a separate tyrosine kinase called JAK2.

EPO-EPO-R pathway:

1. EPO binds to EPO-R on erythroid precursor cells in the bone marrow. This causes the receptor to dimerise/ rearrange.

2. JAK2 is activated. JAK2 is already associated with the intracellular part of EPO-R. Receptor activation brings the JAK2 molecules together. JAK2 phosphorylates itself and then phosphorylates tyrosines on EPO-R.

3. Signalling proteins bind. The phosphorylated tyrosines on EPO-R act as docking sites for signalling proteins.

The major pathway is:

EPO — EPO-R — JAK2 — STAT5 — Gene transcription.