Transforming growth factor Signalling Flashcards
How does TGF-beta induce transcription?
Binds to the receptor -> Causes dimers of type I and type II receptors to obligomerise -> type II receptors phosphorylate and active serine/threonine kinase of type I receptors -> Recruit SMAD -> Type I receptor phosphorylates SMAD -> SMAD4 recruitment -> SMAD dimers form -> Translocate to the nucleus -> Transcription
What is the autocrine hypothesis?
Cancer cells produce TGFalpha and beta and these work on themselves to activate receptors on the cancer cells
What do TGF-beta cytokines regulate?
Regeneration Development Migration Proliferation Immunosuppression Apoptosis Angiogenesis Differentiation
How it is known that type II receptors phosphorylate type I receptors?
Isolate what it is bound to using a radioactive tracer when a fluorescent probe is added
When you remove the kinase activity of type II, you no longer get phosphorylation of type I
But if you remove the kinase activity of type I, this doesn’t effect type II
What is Smad4?
A tumour suppressor
A transcription factor
What is the DNA binding domain of Smad?
Mad homology domain 1 (MH1)
What domain and amino acids are phosphorlyated to recruit SMAD4 to R-smad
SSVS - the second and fourth serine are phosphorylated
What are the 3 groups of SMADs?
R-SMADs (receptor activated SMADs)
Co-mediator SMAD
Inhibitory SMAD
What SMADS are R-SMADs?
Smad1 Smad9 Smad5 Smad3 Smad2
What SMAD is a Co-mediator SMAD?
Smad4
What SMADs are inhibitory SMADs?
Smad6
Smad 7
What domain of SMADs bind to other proteins?
Mad Homology domain 2 (MH2)
What proteins does the MH2 domain bind to?
Receptors and Phosphorylation Homo and hetero-oligomerisation Cytoplasmic anchoring Nuclear import/export Binding to co-activators/ repressors
What Smads do receptor type II and I activate when bound to TGF-beta?
Smad2 and 3
Smad4 downstream of them
After from type I receptor, what other receptor can type II obligomerise with?
ALK1
What Smads do receptor type II and ALK1 activate when bound to TGF-beta?
Smad1, 5 and 8
Smad4 downstream of them
What is the accessory ligand for type I and II receptor bound to TGF-beta?
Betaglycan
What is the ligand trap for type I and II receptor bound to TGF-beta?
LAP
What is the accessory ligand for ALK1 and type II receptor bound to TGF-beta?
Endoglin
Why do cofactors have to bind to the DNA to induce SMAD mediated transcription?
Since they have limited affinity and insufficient selectivity and therefore there would be unwanted transcription if cofactors weren’t needed
What are the context-dependent cellular roles of TGF-beta?
Cell cycle control
Differentiation
Survival
Motility
What biological processes are invoked by TGF-beta?
Haematopoiesis Epithelial to mesenchymal transition Angiogenesis Tissue maintenance, healing and repair Bone formation ES cell pluripotency/ differentiation
What disorders are associated with TGF-beta/BMP ligands?
Myostatin - mutations cause ‘double muscle’
Noggin - mutations cause proximal symphalangism
GDF5 - mutations cause chondrodysplasia
AMH - mutations cause persistent muellerian duct syndrome (development of a uterus in a male)
What disorders are associated with TGF-beta receptors?
ENG, ALK1, BMPRII mutations - hereditary hemorrhagic telangiectasia (HHT)
ALK2 mutations - Fibrodysplasia ossificans progressiva (FOP)
ALK5 mutations - Self-healing aggressive squamous carcinoma
ALK5 and TGFRII mutations - Loeys-Dietz Syndrome
What is hereditary hemorrhagic telangiectasia (HHT)?
Abnormal blood vessel formation in the skin, mucous membranes and organs such as lungs, liver and brain
What is Loeys-Dietz Syndrome?
A connective tissue disorder and therefore can weaken layers of the aorta
What disorders are associated with SMAD mutations?
SMAD4 - Juvenile polyposis and pancreatic and CRC
SMAD2/3 - CRC
What human diseases are associated with impaired BMP/TGFbeta signalling?
Cancer and metastasis Fibrosis Immune disorders Osteoporosis Hereditary spastic paraplegia Arthritis Atherosclerosis Alzheimer's and Parkinson's
What are accessory ligands?
Proteins which bind to the ligands to improve the affinity of the ligands to the receptors
What are ligand traps?
Secreted molecules which bind potently to the ligands to limit the access of the ligands to the receptors
What regulates TGF-beta signalling?
Accessory ligands and ligand traps
Receptor activity and stability
SMAD activity and stability
Transcriptional coregulators/ epigenetic status
How is TGFbeta signalling controlled by the receptors?
When there is too much signalling, TGF-beta initiates SMAD7 and E3 ligase SMURF1 -> Ubiquitylation and degradation of the receptor
When signalling is needed, deubiquitylating enzymes USP4/11/15 are expressed -> deubiquitylation of the receptor -> receptor stabilisation
How do SMADs regulate TGFbeta signalling?
When there is too much signalling, CDKs 8/9 will phosphorylate the SMADs in the linker region and recruit SMURF E3 Ub ligase to ubiquitylate SMAD and remove it from the transcription site
When there is not enough signalling, SCP1/2 will dephosphorylate SMAD at the linker region and PPM1A will dephosphorylate SMAD at the C-terminal so it moves back into the cytoplasm to interact with the receptor again
What prevents SMAD2/3 ubiquitylaton?
OTUB1 (deubiquitylating enzyme)
How is TGFbeta signalling controlled by transcription?
Self-enabling (positive)
Self-enabling (negative)
Switch enhancer
Derepression
Give an example of a self-enabling (positive)
Smad2/4 bind to Foxh1 -> recruited to the nodal response element -> Mix transcription -> Mix recruits Smad complexes to the Mix promoters -> induce target genes e.g. goosecoid
Give an example of a switch enhancer
In ESCs:
Low levels of TSO complex (TEAD, SMAD and OCT4) -> increase pluripotency genes
High levels of TSO complex -> inhibition of pluripotency genes (‘Buffering’)
When the TSO complex is close to FOXH1, it inhibits FOXH1 and therefore inhibits mesendodermal gene transcription
When cells switch to differentiation, the TSO complex is disrupted and SMAD complexes are activated, interact with FOXH1 -> induce mesendodermal genes
Give an example of a self-enabling (negative)
TGF-beta causes SMAD3/4 recruitment to promoter -> induce ATF3 transcription -> ATF3 recruits SMAD3/4 onto a different promoter -> inhibit ID1 expression
Give an example of derepression
A repressor and SKIL/SKI bind to SMAD4 at the SMAD binding element (SBE) to inhibit SMAD3-dependent genes -> TGF-beta signalling -> RNF111 is recruited to SKI/SKIL via activated SMAD2/3 -> rapid degradation of SKI/SKIL -> exposure of SBE -> formation of SMAD3/4 complexes -> SMAD3-dependent genes
