Cell signaling RTK

Define enzyme coupled receptors (+Typical structure)

• Large, heterogeneous family of transmembrane proteins that function as enzymes

• “Single-pass” membrane proteins (typically):

  • Ligand-binding domain on the extracellular side of the PM.

  • Catalytic site on the cytoplasmic side of the PM.

• Typically, act as protein kinases that phosphorylate specific sets of proteins in the target cell.

• Receptor tyrosine kinases (RTKs) are most common class of these proteins

Examples of extracellular signal proteins that act via RTKs

• EGF - epidermal growth factor

• NGF - Nerve growth factor

• PDGF - platelet-derived growth factor

• FGF - fibroblast growth factor

• VEGF - vascular endothelial growth factor

Describe RTK activation

1. The signal protein (ligand) binds to two RTK monomers, bringing them together to form a dimer

2. Closeness of dimers leads to 2 kinase domain phosphorylating e/o. Effects:

   1. at some tyrosines, phosphorylation promotes complete activation of the domains

   2. phosphorylation at other tyrosines generates docking sites for intracellular signaling proteins — results in formation of large signaling complexes; can relay signal downstream

Where do intracellular proteins bind after receptor activation, how are they activated?

• intracellular signaling proteins bind to a particular phosphorylated site on the activate receptors. Activation:

  • once bound to RTK, signaling protein may be phosphorylated on tyrosines → activated

  • binding to RTK alone induces conf. change or brings it near next protein in signaling pathway → activated

Exceptions to typical RTK activation

• insulin receptor

• epidermal growth factor receptor

insulin receptor activation

• receptor is ALWAYS a dimer

• ligand binding causes a conf. change that brings the 2 internal kinase domains closer together

  • there is still autophosphorylation of tyrosines in kinase active sites

epidermal growth factor receptor activation

receptor kinase domain is NOT activated by transautophorphorylation

1. dimerization orients internal kinase domains in an asymmetric dimer

2. activator pushes against receiver

3. conformational change in receiver

4. → phosphorylates both receptors

Describe Ras activation

1. adaptor protein (ex. Grb2) recognizes a specific phosphorylated tyrosine on the activated RTK (by SH2 domain)

2. adaptor protein binds to Ras GEF (ex. Sos) (interaction b/t SH3 domain(s) and proline-rich region of Sos)

3. Ras GEF domain stimulates inactive Ras protein to replace its bound GDP with GTP → Ras activated to relay the signal downstream

Are Ras activations long or short lived? What deactivates Ras?

Tyrosine phosphorylations and Ras triggered by activated RTKs are short-lived

• tyrosine-specific protein phosphotases - reverse phosphorylations

• Ras GAPs - induce Ras to inactivate itself by hydrolyzing bound GTP to GDP

Describe MAP kinase pathway

1. Activated Ras protein recruits Raf (MAP kinase kinase kinase) to the plasma membrane and helps activate it

2. Raf activates the MAP kinase kinase Mek by phosphorylation

3. Mek activates the MAP kinase Erk by phosphorylation

4. Erk phosphorylates many downstream proteins, incl:

   1. protein kinases (changes in protein activity)

   2. transcription regulatory proteins in the nucleus (changes in gene expression)

How is the MAP kinase pathway regulated?

through positive and negative feedback loops (*see slides and txtbk*)

proto-oncogenes

genes that normally control how often a cell divides and the degree to which it differentiates or specializes

oncogenes

permanently “turned on” or activated gene when it is not supposed to be; accelerate cell growth and division

where are proto-oncogenes found in normal cell growth pathway?

extracellular growth factor binds to receptor → activates signaling enzymes → activates transcription factors in the nucleus → proto-oncogenes control cell proliferation

Ras oncogenes

point mutations create a hyperactive Ras (GTPase) that can not shut itself off by hydrolyzing bound GTP to GDP

• constant signaling for cell growth and proliferation

• the oncogenes stimulate growth even when there is no growth factor produced

PI-3 Kinase activation of phosphatidylinositols (PI)

the plasma membrane bound enzyme phosphoinositide 3-kinase (PI 3-kinase) phosphorylates the inositol ring on phosphoinositide lipids.

• after 3 phosphorylations, PI(3,4,5)P3 is produced, which serves as a docking site for intracellular signaling proteins