L2 - How are promoters selected? Flashcards
How is DNA packaged?
146 bp of DNA wraps round histone forming nucleosomes
What feature do some promoters have to keep txn?
Sequences that discourage nucleosome occupancy
How is txn inhbited?
methylation at the CpG
What is the cpG
Regions enriched for CG dinucleotides (CpG islands) with promoters in
What happens when CpG is unmethylated?
Recruits TFs such as CFP1 and MLL family
What do CFP1 and MLL do?
Recruit H3K4 methyltransferase complexes and allow it to spread to adjacent nucelosomes
What do H3K4Me3’s do?
Trimethlate lysine 4 of histone H3
What can mutations in MLL cause?
Aggressive leukaemias resulting from fusion proteins misregulating target genes
What recognises H3K4me3 and what does it do?
Chromatin remodeller (CHD1) Helps move nucleosomes away from promoter
What subunit of TFIID also recognises H3K4me3?
Complex of TBP with 13 TAFs (TBP associated factor) (TAF3 binds H3k4me3)
How was it found which proteins and subunits bind to each other?
Pull down assays with beads
What is ChIP and what is used for?
Chromatin immunoprecipitation
To test which regions of DNA are bound by a protein in cells
What parts of TFIID contribute to promoter recognition?
TAF1 - binds histone H4 and initiator (Inr) DNA
TAF6 - binds downstream promoter element (DPE)
TBP binds TATA box
Why is there so many sites of contact for TFIID?
Provides versatility for different promoter arrangments (TATA-less promoters)
How do TFs help initation?
Assist in recruitment of TFIID, basal factors and pol 2.
Very inefficient with out help
What is an example TF helping in recruitment at pol 2?
Sp1 binds g/c rich sites in DNA of many promoters and recruits TFIID with TAFs
What do activating TFs also do?
Recruit chromatin remodellers and histone modifiers
What does p53 recruit other than TFIID?
p300 histone acetyltransferase (HAT)
What does HAT do
acetylates histone H4 so it is recognised by TAF1
What are LOF mutations in chromatin remodellers associated with and exmaple
Various cancers
SNF5 deletions in malignant rhabdoid tumours
What did ChIP-PCR show about histone H4?
Levels incease when p53 and p300 bind a gene induced by UV irradiation
What are the erasers of acetylation on histones?
HDACs (histone deacetylases)
What recognises acetylated histones?
Bromodomains
What does TAF1 having 2 bromodomains do?
Allows simultaneous recognition of two close acetylated lysines giving enhanced specificity
What can be used to treat cancers?
HDAC inhibitors to reactive silenced genes
What is an example of an HDAC inhibitor
SAHA
what do activating TFs also recruit?
Basal TFs directly and indirectly
What can pioneer TFs do? examples
Bind nucleosomal DNA and increase accessibility
e.g. FOXA1, AP1
What does binding of AP1 to DNA do?
Allows access of glucocorticoid receptor to adjacent site - allowing cooperative binding of other TFs
What is interesting about FOXA1 and what does it allow?
DNA binding domain resembles linker histones
Allows oestrogen receptor (ER) to access DNA
Binds enhancers, not promoters
How is FOXA1 implicated in breast cancer?
Overexpress ER, tumour growth is hormone sensitive
FOXA1 or AP1 may redirect ER to alternative gene targets increasing severity
What are the features of enhancerS?
Clusters of TF binding sites
1m in human genome
Can be 100s of kb from gene
Marked by histone acetylation, DNase hypersensivity and H3K4me1 (different from 3)
What is chromosome conformation capture use for?
Test interactions between regions of DNA in cells
What feature do enhancers have?
Cell type selective
How are chromatin loops stabilised?
Cohesin and mediator
What are the features of cohesion?
Rig shaped to accommodate 2 strands
Stabilise interactions between enhancer and promoter
Mutations result in developmental defects (retardation)
What are the features of the mediator?
26 subunits
Binds unphos CTD of pol 2
Target for many activators
Mutations result in developmental and neurological disorders
