Ser/Thr kinase receptors

What are TGF-β receptors?

• All members of the transforming growth factor-β (TGF-β) receptor family have intrinsic Ser/Thr (ST) kinase activity.

• They are involved in many cell processes, such as development and homeostasis.

How are TGF-β receptors activated?

• The Type II receptor is a constitutively active dimer.

• The Type I receptor (also a dimer) is the one that is switched on.

–    Ligand binds → the Type II receptor recruits and phosphorylates the Type I receptor on Ser/Thr.

–    Type I and Type II form a heterotetramer.

–    Phosphorylated Type I recruits and phosphorylates R-SMAD.

–    R-SMAD dissociates from the receptor and binds Co-SMAD → the SMAD complex.

–    The SMAD complex translocates to the nucleus and binds the SMAD binding element (SBE) in target genes → regulates transcription.

How does phosphorylation activate the Type I receptor?

• Type I kinase activity is autoinhibited by its GS domain (glycine/serine-rich).

• The Type II receptor phosphorylates the GS domain on TTSGSGSGLP.

• This causes a conformational shift, pulling the inhibitory loop out of the active site.

What are the types of SMAD proteins?

There are 8 SMAD proteins, in 3 groups:

• R-SMAD (receptor SMAD): SMAD 1, 2, 3, 5, 8. Different receptors (e.g. TGF-βR) bind different R-SMAD.

• Co-SMAD: SMAD 4. Common to all receptors.

• I-SMAD (inhibitory SMAD): SMAD 6, 7.

What is the structure of SMAD?

• MH1 (Mad homology 1) → linker → MH2 (Mad homology 2) → SSxS (serine-rich) C-terminal motif.

• Basal state: MH1 and MH2 bind to each other to form an autoinhibitory domain (AID). MH2 can also bind the SARA adaptor.

• Activated state:

–    MH1 contains an NLS and can bind DNA

–    MH2 interacts with the receptor and with DNA-binding proteins, mediates the R-SMAD–SMAD4 interaction, and can activate transcription

What are the regulatory phosphorylations of SMAD?

• C-terminal Ser-Ser-X-Ser (end of MH2) is phosphorylated by the Type I TGF-βR → activation.

• MAPK phosphorylation of the linker → inhibition of SMAD.

How do SMAD proteins link to TGF-β signalling?

• R-SMADs are brought to the membrane by SARA (SMAD anchor for receptor activation), which binds membranes via its FYVE lipid-binding domain.

–    TGF-β induces receptor oligomerisation, and the Type II receptor phosphorylates the Type I receptor.

–    The R-SMAD–SARA complex is recruited to the Type I receptor via the MH2 domain of R-SMAD.

–    Type I phosphorylates R-SMAD on its C-terminal tail (serine-rich domain).

–    R-SMAD dissociates from the receptor complex, binds SMAD4, and translocates to the nucleus to regulate transcription.

How does I-SMAD regulate the pathway?

• I-SMAD competes with SMAD4 for binding to R-SMAD.

• I-SMAD also binds the activated receptor and recruits the SMURF ubiquitin ligase, causing receptor degradation.

• I-SMAD sets the duration of TGF-β signalling by negative feedback. It is a downstream target of the R-SMAD–SMAD4 complex, which activates it; I-SMAD then inhibits the pathway.

How does RAS-ERK signalling integrate with TGF-β signalling?

RAS-ERK signalling inhibits TGF-β signalling. Activated RAS → ERK, which phosphorylates the linker between MH1 and MH2 of R-SMAD → blocks nuclear translocation of the SMAD complex.

How does TGF-β cause epithelial-mesenchymal transition (EMT)?

• TGF-β induces EMT, which contributes to development, tissue fibrosis, wound healing and cancer.

• EMT is characterised by:

–    Loss of intercellular junctions and polarity.

–    Increased motility.

–    Loss of expression of epithelial proteins and increased expression of mesenchymal proteins.

What genes are altered during EMT?

• SMAD signalling recruits:

–    Epithelial repressors: which represses epithelial genes: E-cadherin, claudin, occludin.

–    Mesenchymal activators: which promotes mesenchymal genes: vimentin, α-SMA, PAI-1, α2 collagen.