Wnt Beta-Catenin/ TCF4 pathway Flashcards
FAP
- name and what it predisposes to
= Familial Adenomatous Polyposis
- inherited disorder
Predisposes to colorectal cancer, small bowel cancer + stomach cancer
FAP
- allele type
Autosomal dominant
= 1 copy of mutant gene
= disease
AFAP
Attenuated version of FAP
- lower risk fo colorectal cancer
FAP
- mutated gene
APC gene
APC
- name
Adenomatous Poliposis Coli
APC
- inactivated by
- where mutations occur
Mutations
LOH
Promoter hypermethylation
Mutation cluster region
Colonic crypt
- what is it?
All stem cells in small intestine reside
Colonic crypt
- what happens?
Epithelial renewal occurs:
- Proliferation
- Differentiation at mid-crypt
- Migration towards intestinal lumen
- Pluripotent stem cells at bottom of crypt generate progenitors
Colonic crypt
- Beta-catenin + APC
Mutation in Beta-catenin or APC in differentiated cells with normally arrested cell cycle
- > progenitor-like phenotype
- > accumulation at site of future polyp formation
Beta-catenin
- role
Key molecule
- translocates to nucleus to activate gene transcription upon Wnt activation
Beta-catenin
- activates transcription
Binds to LEF-1/TCF4
- > activates transcription of Wnt target genes
- > block differentiation + promote proliferation
Beta-catenin
- stability
Regulated by destruction complex made up of proteins including axis + APC
Causes of increased beta-catenin in nucleus
- mutations
APC mutation
- prevents recognition of beta-catenin
Beta-catenin mutation
- prevents phosphorylation + associated destruction APC complex
Lead to more beta-catenin in nucleus
-> increased transcription
Wnt-Signalling ON
- Wnt ligand binds to receptor
- Inactivation of APC
- Beta-catenin accumulates
- Translocates to nucleus
- Activates genes that promote cell proliferation
Wnt-Signalling OFF
- Wnt ligands prevented from binding
- APC recruits other components
- Degrades beta-catenin
- No proliferating genes transcribed
DKO
- what are they?
= knock-out cells
derived from HCT116 lacking the DNA methyltransferases (DMNT)
(=they can’t methylate DNA)
DAC
- name + role
5’-aza-2’-deoxycytidine
Inhibits DMNT activity
Trichostatin A
- role
Inhibits histone deacetlyase activity
Cells treated with DAC
Inhibits DMNTs
- > decreases Sox17 methylation
- > increases Sox17 expression
Cells treated with DAC inhibitor
- conclusion
DMNT can methylate promoter
-> Sox17 not expressed
Soz17 is hypermethylated in CRC cells
Methylated-specific PCR
MSP
1 set of primers designed to amplify methylated DNA (M) and 1 set designed to amplify unmethylated DNA (U)
CpG islands
- when are they methylated?
Control tissues unmethylated
Sox17 methylated in cancer
Bisulfite genomic DNA sequencing
- what is it?
Provides added info compared to MSP
- can determine methylation status of individual Cytosines by sequencing
Bisulfite genomic DNA sequencing
- DKO of DNMT1 + 3b
- Beta-catenin activating mutation
- APC inactivating mutation
Mostly unmethylated
Heavily methylated
Heavily methylated
