L7 - Txnal Regulation of oncogenes and tumour suppressors Flashcards
What does inheritance of one inactive RB allele give?
90% chance of developing retinoblastoma at any early age after somatic mutation of second allele
What is mutation of RB gene also common in?
Many tumour types
Almost all small cell lung cancers
In cervical cancers RB neutralised by E7 from HPV that binds to the pocket of RB
What is the structure of the RB pocket?
Two subdomains each resembling a cyclin fold that interact to form a single domain
What is the structure of the rest of RB?
N terminal domain has cyclin folds
Rest unstructured and flexible
C terminal bound by protein phosphatase 1 and CDK 2,4,6
What happens to RB in G1 phase?
Hypophosphorylated by cdk4 and cdk6 +cyclin D committing cell to leave G1 (restriction point)
What is the function of cyclin D in the cell cycle/
Expressed in response to mitogens - RB therefore couples cell cycle entry to mitogenic signals (gatekeeper)
What happens to RB in S phase?
cdk2/cyclin E phosphorylate further and inactivate RB
What happens to RB in G2?
cdk2/cyclin A maintains phosphorylation and inactivation
What happens to RB in M phase?
Phosphorylation reversed by protein phosphatase 1 (PP1)
What happens when RB is hypophosphorylated?
cells remain in G1 or withdraw into a quiescent state
How can some cancers subvert Rb?
Neutralise by binding of oncogenic protein to pocket domain or switched off by phosphorylation
How can cancers hypophosphorylate RB?
Overespress Cyclin D and ckd4
What is the function of INK4?
Encodes p16 and p15 that bind to cdk4 and 6 blocking action so cant bind to cyclin D
How is INK4 involved in cancer?
When INK4 genes deleted (melanoma) this deregulates RB phosphorylation
What TFs does RB regulate?
MyoD, UBF, TFIIIB, E2Fs
What are E2Fs involved in
Regulate DNA syn genes and cyclin E gene
Recruits TFIID to promoters
How does RB regulate E2Fs?
Binds txn action domain
RB recriots HDACs and chromatin remodellers to repress E2F
Phosphor of RB causes release
What do activator E2Fs do?
Txn activators during cell cycle progression and most abundant in proliferating cells
What do repressor E2Fs do?
Expressed in quiescent cells, inhibit E2F target genes
How is RB involved in pol 3 txn?
Hypophos RB binds to TFIIIB and blocks interactions with C + pol 3
So RB can block all pol 3 txn
What other proteins have pocket domains?
p107 and p130 (54% identical to each other and 25% identical to RB)
What is a characteristic of p107 and p130 and how is this involved in cancers?
Can be switched off through phosphor
So a lot of cancers lose function of all 3
What is ARF>
E2F target
One of 3 tumour sup genes in a 35kb region often deleted in tumours
How does ARF work?
Induced by oncogenic signals
Binds MDM2 and inhibits ubiquitin ligase activity
Triggers p53 response
What does pol 1 do?
Syntehsise 18S, 5.8S and 28S rRNA as large precursor transcript
How does ARF affect the activity of pol 1?
Inhibits processing of S’s rRNA
What are free ribosomal proteins and example of one?
Released by imbalances in ribosome biogenesis (nucleolar stress) eg RPL11
What can RPL11 do?
Bind and inhibit MDM2 - activating p53
What does activating p53 do during nucleolar stress?
Provides important checkpoint to prevent cell cycle progression when ribosome production is inaequate
What are p53super mice and what did they show?
Extra copy of p53 inserted in
Reduced cancer incidence
What does loss of RB function cause in cancers?
Induction of ARF and hence p53 leading to oncogenic stress
How does HPV cause cancer?
Encodes oncogenincE7 that binds and neutralises and E6 targets p53 for ubiquitination and degradation - causing cervical cancer
How does SV40 DNA tumour virus cause cancer?
Large T antigen binds and neutralises RB and p53
What is the incidence of loss of p53 in all cancers?
Over 50% (highest of all known genes)
Where is p53 usually mutated?
DNA binding domain
What do MDM2 and MDMX do?
Bind txn domain of p53 (mono export, poly degrade)
Both bind and inhibit RB
What do intentional mutations of p53 in mice show?
Knock in - prevents induction of apoptosis but not tumour suppression
p53 mutants able to induce cell cycle arrest and apoptosis, but not supress tumour
What occurs in Brukitt lymphoma?
Chromosomal translocations place MYC under control of an Ig enhancer
How does MYC generally act?
Stimulates txn of open genes rather than inducing silent ones
How does the way MYC usually acts cause cancer?
May amplify other oncogenic TFs
contrasts normal assumption that oncogenic transformation involves switching key genes or or off
How did they show what genes MYC activated and repressed?
Model cell line carrying MYC transgene controlled by a doxycycline-activated promoter
What genes did they show MYC activated and repressed?
Increased pol2 occupancy at KLF15
Reduced pol 2 occupancy at ALDJ3B1
How can repression by MYC sometimes work and give an example?
Involving interference with other activators
Miz1 activates p21, but MYC blocks interaction with p300
How is MYC involved with pol1?
DNA sites recognised by MYC found in rRNA promoters, binds and stimulates txn but recruiting SL1 and HATS
How is MYC involved with pol3 txn?
MYC recruited by TFIIIB where it then recruits HATs to stimulate txn
What does the action of MYC have to do with cancer?
Involved in all protein synthesis required for tumour growth
How did they show MYC induced increased protein synthesis?
Used MYC overexpresser in mice B cells, stimulated protein syn and size of b cells
Then crossed with a lower translating mutant (L24) and reverted to normal (control)
What was a therapeutic option for MYC driven cancers?
L24 decreased lymphomagenesis suggesting MYC needs to increase rate of protein syn to cause lymphomas
