Tumour Suppressor Genes Flashcards
1
Q
what is the function of TSG
A
The protein products of TSGs function to negatively regulate cell growth, and/or are components of cell cycle checkpoints that ensure genomic integrity in response to genotoxic stress.
- e.g. in response to DS DNA break induced by ionising radiation, cell cycle check point will be invoked → cell cycle halted → DNA repaired before cell cycle continues.
- If the damage is too great, apoptosis will be activated.
2
Q
what happens if TSG product function is lost, how does this happen, and what does this result in
A
- Loss of TSG product function means loss of growth suppressive function and/or loss of cell cycle checkpoint control.
- occurs through gene mutation
- Results in enhanced and dysregulated proliferation
3
Q
what is required for cancer to occur
A
Cancer is a multi-step process – requires activation of many oncogenes and inactivation of many tumour suppressor genes.
4
Q
are TSGs dominant or recessive in cancer
A
TSGs are recessive in cancer. Both alleles (inherited from Mother and Father) of a TSG must be eliminated or inactivated to abolish Tumour Suppressor properties.
5
Q
can TSG mutations be heritable
A
TSG are heritable if mutation occurs in the germline (sex cells)
They are somatic if mutation occurs only in the cell where the tumour originates.
6
Q
what is the 2 hit hypothesis
A
Known as Knudson’s two-hit hypothesis – both alleles need to be eliminated for cancer to occur.
7
Q
characteristics of inherited form of cancer
A
- Alfred noticed that in Inherited form of cancer:
- One mutation in germline (sex cell)
- Second mutation was somatic – occurred in cell where cancer originated.
8
Q
characteristics of Non-hereditary form of cancer
A
Both mutations are somatic and occurred specifically in cell where cancer originated.
9
Q
inherited vs non-inherited forms of cancer - which one is associated with increased cancer susceptibility and why
A
If TSG mutations occur in sex cells the individual formed from, the person will have the mutation in every cell of their body.
Therefore, these syndromes are associated with increased cancer susceptibility
10
Q
which form of cancer is more common
A
The vast majority of human cancers (90%+) are somatic and are associated with mutations of TSG only in the cell where the cancer originates – they are non-heritable.
11
Q
TSGs: inherited syndrome and cancer susceptibility
A
SEE IMAGE
12
Q
what is the normal role of RB1 and TP53
A
normal role of RB1 and TP53 gene products in cell growth and cell death
13
Q
what is retinoblastoma
A
- it is cancer of the retina
- Picture shows longitudinal section of eye with retinoblastoma tumour displacing retina and growing into the vitreous humor.
- We can also see thickening of the optic nerve due to the metastasis of the tumour
14
Q
germline mutation in which gene gives rise to retinoblastoma
A
Germline mutation in RB1 gives rise to retinoblastoma which can result in both retina and bone cancers. (retinoblastomas + osteosarcoma)
15
Q
Familial v Somatic Retinoblastoma
A
- In Familia retinoblastoma the fertilized egg possesses 1 mutant RB allele and 1 normal (wild type) RB allele.
- In the familial form of retinoblastoma every cell in the body possesses one wild type RB allele and 1 mutant RB allele.
- After the first somatic mutation of RB gene in the retinal cell from which the cancer originates from, there is now 2 mutant RB alleles.
- Therefore there is inactivation of its tumour suppressor properties
- In somatic retinoblastoma both alleles are normal (wild type).
- So in somatic retinoblastoma the first somatic mutation results in 1 mutant and 1 wild type allele.
- A second somatic mutation in retinal cell where cancer originates is required to inactivate the tumour suppressor properties of RB.
16
Q
which form of retinoblastoma shows earlier onset and why
A
- familial
- As familial retinoblastoma only requires 1 somatic mutation in RB compared to 2 in somatic, it presents earlier than somatic retinoblastoma
17
Q
what differentiates familial and somatic retinoblastoma (presentation-wise, other than onset time)
A
- familial is characterised by bilateral disease – patient has retinal cancer in both eyes.
- Whereas somatic retinoblastoma the patient has unilateral disease – cancer is in one eye.
18
Q
Retinoblastoma inheritance pattern
A
- facial/inherited form
- We have an unaffected female and affected male in first generation which give rise to daughter who carries mutant RB allele and presents with retinoblastoma.
- In 3rd gen she passes on mutant RB allele to a male and female and also has unaffected male and female.
- In this scenario the inherited form of retinoblastoma patients presents with:
- Multiple tumours that are often bilateral and early onset.
- They also have an Increased cancer risk, particularly soft tissue sarcomas, and osteogenic sarcomas
- In the somatic form
- There are no affected individuals in early generations however in 3rd generation we have a case appear which indicated by looking at pedigree that this mutation is likely to be somatic.
- In this scenario the somatic form of retinoblastoma patients presents with:
- Single tumours that are often unilateral and late onset.
- They also have an Increased cancer such as sarcomas
19
Q
when can the familial/inherited pattern of retinoblastoma look like a somatic form, when in fact it is a inherited form.
A
- sometimes mutations can occur in germline in further generations
- In this case it will look like a somatic form when in fact it is a inherited form.
- This is known as a de novo familial retinoblastoma.
20
Q
which non-retinal tumours are familial retinoblastoma patients most susceptible to
A
- Those individuals who have bilateral (familial) disease have increased susceptibly to non-retinal tumours such as osteosarcomas during their lifetime compared to those who have unilateral disease.
- This is represented on graph - 1/3 chance of having non-retinal tumour for bilateral disease compared to 6% for unilateral disease.
21
Q
how do childhood Retinoblastoma patients present (2)
A
In childhood retinoblastoma, patients often present with either white light reflection and or a squint.
22
Q
what type of mutations do we see in familial retinoblastoma (3 )
A
- In Familial retinoblastoma, the mutations we see:
- large deletions (about 20%);
- single base substitutions (about 50%)
- small length mutations (about 30%)
- These affect expression of RB gene and changes amino acid statuses – which affect function of protein produced.
23
Q
what type of mutations do we see in somatic retinoblastoma (3)
A
- In Somatic retinoblastoma, the mutations we see:
- Gene deletions, base substitutions, small length mutations
24
Q
complete vs incomplete penetrance
A
- Most mutations are associated with almost complete penetrance because it is an autosomal dominant disease.
- Rare alleles show incomplete penetrance and reduced expressivity – known as low penetrance retinoblastoma.
25
how do we lose the second wild (normal) type of allele RB (2)
- Familial and Somatic – how do we lose second wild type allele of RB
- Often 2nd independent mutation (~30%) and a small amount of hypermethylation of the 5’ region- RB1 gene
- Approx 70% of 2nd mutation are through loss of heterozygosity during mitotic recombination; mitotic non-disjunction; large deletions
26
are TSGs recessive or dominant at the genetic level
- TSGs are recessive at the genetic level:
- Both alleles need to be inactivated to inactivate the tumour suppressor properties of the protein.
27
are TSGs recessive or dominant at the cellular level
- TSGs (e.g. *RB1*) are often dominant at the cellular level.
- An Autosome indicates that the gene locus is on a non-sex-chromosome
28
how is the second mutation usually acquired (in people with inherited mutation) (3)
- i.e. if you inherit one mutant allele, you have a high probability during your lifetime of acquiring a second inactivating mutation in that gene, usually through:
- loss of heterozygosity
- 2nd independent mutation
- promoter methylation
29
How does loss of heterozygosity occur at cellular level
- In an affected individual in G1 phase we have one chromosome with wild type RB allele and 1 mutant RB allele
- Following DNA replication in S phase we now have 2 copies of wild type allele and 2 copies of the mutant allele.
- During G2 and mitotic phases of cell cycle chromosomes undergo homologous recombination that facilitates exchange of genetic loci between chromosomes.
- Upon chromosome segregation at anaphase onset in mitosis we produce daughter cells that have either retained Rb heterozygosity (1 wild and 1 mutated allele) or they lose heterozygosity (daughter cell has 2 wild types or 2 mutant type alleles)
- The daughter cell with the 2 mutant type alleles is where the cancer originates as RB as been inactivated and cell will undergo uncontrolled proliferation.
30
on which chromosome is the RB locus in normal vs retinoblastoma
chromosome 13
- In these chromosome banding images banding is lost as 13q1 near centromere.
- This is shown schematically below that.
- Normal chromosome you have genetic locus but in retinoblastoma chromosome you have lost the locus
31
what is the gene product of RB1
- Retinoblastoma gene: RB1
- Retinoblastoma gene product (protein): pRB
32
what is the role of the Retinoblastoma protein in cell cycle control
- What is the normal function of pRB?
- pRB is hypo-phosphorylated in G0 and early stages of G1 - acts as. restriction point.
- Whereas as the cell progresses through G1 and passes the restriction point pRB becomes increasingly phosphorylated and eventually becomes hyper-phosphorylated such that cells can pass through the restriction point and into s phase.
33
what is the function cyclin/Cdk in cell cycle
We know that cyclin/Cdk families of proteins drive cell cycle progression.
- Cyclin D/CDK46 complexes that function in early G1 up to restriction point are responsible for driving phosphorylation of RB at this time.
- From the restriction point onwards into S phase cyclin E CDK2 is active and this drives the phosphorylation of RB at this time.
34
what is the function of pRB
- It functions as a transcriptional repressor and it forms large repressor complexes during G0 and early stages of G1 such that it inhibits E2FDP regulated promoters to prevent cell cycle progression and the transcription of those genes important for cell cycle progression.
- However, upon phosphorylation of the pRB protein by cyclinD-cdk46, this allows the repression complex to change it its character such that histone DAC is removed from pRB which allows this p300 to prime some e2f responsive promoters for transcriptional activation.
- Upon cyclin e cdk2 phosphorylation of RB – it dissociated from ESFDP transcription factors which allows E2fDP to drive transcription of those genes required for passage of cells through restriction point and entry into S phase.
35
what happens to pRB in cancer
pRB is inactivated/mutated in human cancer
36
what does the inactivation of pRB in human cancer lead to
pRB inactivation in human cancer leads to bypass of restriction point and entry into S-phase of cell cycle.
- This leads to bypass of restriction point and entry into S phase.
- In normal cells pRB functions to repress e2fDP regulated transcription so it is a regulated phenomenon that is controlled by pRB phosphorylation.
- In cancer we lose RB so E2F DP transcription factors are now constitutively active.
- So they bypass the restriction point and you get uncontrolled entry into S phase and progression through cell cycle.
37
what regulates pRB and how
Regulation of pRB by the p16ink4a Cdk inhibitor
Cyclin Cdks are regulated themselves by Cdk inhibitors such as p16ink4a inhibitor which can prevent CDK dependent phosphorylation of pRB.
38
describe what happens in situations of cellular stress
in situations of cellular stress, p16ink4a expression is increased which inhibits CDK4 kinase activity towards pRB such that pRB forms this transcription repressor complex on E2F and promotes cellular growth arrest or cellular senescence which is a form of terminal grow arrest.
39
what happens to p16ink4a in cancer
p16ink4a is a TSG product inactivated in human cancer - particularly melanoma
40
how is p16ink4a inhibited
P16ink4a is inhibited through mutation and epigenetic silencing (e.g. promoter methylation)
41
what are the effects of P16ink4a inhibition
if you lose p16ink4a then we inhibit the regulation Cyclin D Cdk4/6 which can then phosphorylate pRB and allow for passage through the restriction point and entry into S phase.
42
what is the gene product for TP53
- Gene: *TP53*
- Gene product (protein): p53
43
what is the function of p53
p53: ‘Guardian of the genome’ – protects cells against genetic instability in response to genotoxic and cellular stresses.
44
how many human cancers carry mutation in TP53 gene
- Between 30% and 50% of all human cancers carry missense mutations in *TP53* gene that give rise to mutant p53 protein.
- It has been Suggested that p53 pathway inactivated in most (over 90%) of all human cancers
- i.e. p53 function is inhibited, although *TP53* it is not mutated (the p53 pathway is inactivated in retinoblastoma)
- This means that those proteins that lie both upstream and downstream of p53 activation might also be mutated at the gene level such that they are inactive.
- Therefore even if you have a wild type p53 protein its affected is still supressed due to mutated inactive proteins that lie upstream/downstream of its activation
45
what syndromes do germline mutations in TP53 result in
Li-Fraumeni and Li-Fraumeni-Like Syndromes
46
are these syndromes heterogeneous or homogenous
This is a clinically and genetically heterogeneous inherited cancer syndrome
47
is it dominant or recessive
It is autosomal dominant – acquire one mutation through the germline then through loss of heterozygosity you get mutation of the second allele.
48
what characterises mutations in TP53 (3)
early onset of tumours
multiple primary tumours within an individual
multiple affected family member.
49
Li-Fraumeni gives rise to Multiple tumour types, name them (5)
- soft tissue sarcomas and osteosarcomas,
- breast cancer,
- brain tumours,
- leukaemias,
- adrenocortical carcinomas
50
how prevalent are TP53 somatic mutation in human cancer
- TP53 mutation is also associated with somatic human cancer. That is cancers that occur through mutation of the TP53 gene is the actual cell where the cancer originates.
- These are non-heritable cancers.
- In almost every cancer p53 is mutated to an extent.
51
when is TP53 typically inactivated
TP53 is typically inactivated during the late stages of cancer progression.
52
what are the functions of p53 (4)
- p53 protein is a sequence-specific transcription factor. It has 4 primary activities:
- Pro-apoptotic activity → can induce cell death
- Growth arresting activity → can transiently arrest the cell in a particular stage of cell cycle.
- Induces cellular senescence → terminal growth arrest phenotype.
- Inhibits angiogenesis → inhibit blood supply to tumour.
53
what happens to mutant p53 (2)
- Mutant p53 loses ability to activate transcription *i.e.* mutant p53 loses growth suppressive properties → uncontrolled growth.
- There are also a class of mutant P53 proteins that have gain of function
- Gain-of-function mutants:
- have oncogenic potential and promote genetic instability
- (And also increase drug resistance)
54
how does the p53-tumour-suppressor-protein function
- A schematic representation of prototypic p53 form is shown below:
- There a number of domains in P53 protein that have specific activity.
- The N-terminal region is involved in transcriptional activation
- Proline-Rich Domain is important in apoptosis
- Sequence-Specific DNA Binding domain is important in sequence specific transcription
- Towards the C terminal region there is:
- A Nuclear localisation signal and an oligomerisation domain (as p53 functions as a tetramer)
- A C terminal regulator domain where p53 function is regulated by post translation modifications such as phosphorylation etc.
- There are 12 p53 isoforms – differing in gene and protein structure.
- This means there are 12 different forms that can be expressed in a cell.
- It is 393 AA in length.
- p53 is a sequence-specific transcription factor.
55
what does p53 activate
- P53 functions a tetramer to activate gene transcription:
- P53 binds to promoter response elements through consensus sequence but this varies between promoters.
- This will drive transcription in collaboration with basal transcription machinery.
56
how is p53 activated
- p53 exists normally as an inactive form in non-stressed cells
- In response to cellular stresses, p53 is activated and the type and extent of damage dictates how p53 activity is modulated to produce the cellular outcome that is necessary.
- In terms of cellular stress:
- dsDNA breaks will activate p53.
- Oncogene activation – increased expression of oncogene products (e.g. Ras) will activate p53.
- Changes in oxygen levels – hypoxia and reactive oxygen species will activate p53
- Changes in proteins translation through the ribosome and loss of cellular adhesion
- These are all cellular stresses
57
how do cellular stresses regulate p53 activity
- It is dependent on type and extent of damage.
- These stresses signal through a class of proteins called mediators e.g. atm & CHK2 and P19ARF etc.
- P53 is normally in its inactivate state.
- But in response to cellular stress ATM and CHK2 are activated and they phosphorylate P53.
- P19ARF modulates MDM2 activity which then all have the effect of activating p53.
- P53 can then work in combination with other proteins such as CBP in complexes.
- These modulate p53 activity.
58
what does p53 do in response to these stressors
- The extent of damage modulates p53 interaction in its post-translation modification states so you get a distinct p53 signatures that when combined in a specific manner to promoters to give the biological response.
- This p53 signature is different between the type and extent of stressor and this is what directs p53 to appropriate promoters.
- P53 can affect anti-angiogenesis through modulation or expression of these genes
- It can induce growth arrest through translation of p21 Cdk inhibitor
- It can promote DNA repair through GADD45
- If DNA damage Is too great then p53 can induce apoptosis through expression of BAX puma and Fas etc.
- P53 also induces expression of Mdm2 which is a e3 ubiquitin ligase which feeds back to inactivate p53 through targeted ubiquitylation in c terminal regulator domain.
59
what is the role of p53 in G1-S checkpoint control/how does p53 function in the cell cycle
- RB is important in passage of cell through restriction point.
- P53 plays an important role in G1-S checkpoint control
- In response to a DNA double strand break induced by e.g. ionising radiation a number of sensing proteins associate with the double stand break and they recruit and activate the ATM kinase.
- The ATM kinase undergoes auto-phosphorylation on serine1981 in response to double strand break formation and it has 3 principle activities:
- Phosphorylate p53 directly on serine 15 to increase transactivation capacity.
- It can phosphorylate the chk2 kinase on threanine68 to activate chk2. Chk2 will then phosphorylate p53 on serine 20. This inhibits the ability of P53 to associate the MDM2 E3 ubiquitin ligase.
- ATM can phosphorylate mdm2 on serine 395 which limits the ability of MDM2 to shuttle pf3 out of the nucleus and target it for degradation in the cytoplasm
- A consequence of this is that p53 is activated and becomes transcriptionally active and drives the expression of Cdk inhibitor p21.
- This is important in cell cycle arrest and p21 can associate with cyclin Cdk partners such as cyclin D-CDK4/6 and cyclin E –cdk2 to inhibit their kinase activity and prevent progression of the cells through G1 into s phase.
- So we put a stop onto this progression.
60
how does p53 induce apoptosis and when (2)
p53 induces apoptosis through both transcriptional and non-transcriptional mechanisms
- When the DNA damage is too great so it cannot be repaired
61
describe how p53 induces apoptosis
- Upon DNA damage p53 is stabilized and accumulate and induces expression of pro-apoptotic molecules e.g. PUMA and BAX which activate the intrinsic pathway of apoptosis by causing mitochondrial outer membrane permeabilization → release of cytochrome → the formation of the azotosome and activation of caspase → which result in cell death.
- P53 through protein interactions can also activate the intrinsic pathway of cell death through binding to anti-apoptotic proteins such as BCL-2 which also results in MOMP and the activation of caspase dependent cell death.
62
describe the p53-Mdm2 auto regulatory feedback loop
- p53-Mdm2 forms an autoregulator feedback loop such that active p53 can induce mdm2 expression.
- The p53 tetramer binds to a response element that lies between exon 1 & 2 of mdm2 gene to drive expression of mdm2 e3 ubiquitin ligase.
- NB: Mdm2 promotes p53 poly-ubiquitylation and degradation once p53 has performed its cellular function
63
what do most of p53 mutations do
- Arginine 248 (red) fits directs into minor grove in DNA.
- So when it mutates, we lose this contact point such that p53 can no longer interact with DNA.
- Other mutations shown here on p53 monomer (pink) also affect p53 association with DNA indirectly – change conformation of p53 so can no longer fit in to the major and minor grooves of DNA
- loss of p53 transcriptional activity in cancer – that is the relationship between p53 mutation, transcription and cancer.
64
what does BRCA1 mutation predispose patients to
- Predisposes to early onset breast carcinoma and ovarian tumours
- Greater incidence of breast v ovarian cancer
- BRCA1 mutation may also be associated with some prostate cancers
65
prevalence of BRCA1 mutations
- Approx 1 in 500 women have germline mutation in BRCA1 gene
- BRCA1 accounts for about 60% of inherited breast cancer
- BRCA1 mutations account for about 5% of all breast cancers and up to 12% of early onset BC
66
important founder mutation
Important founder mutation- 185delAG found in 1% Ashkenazic Jewish population
67
what is the role of BRCA1 in DNA damage checkpoint signalling
- BRCA1 protein has been shown to be involved in the cellular response to DNA damage where it functions in both cell cycle check point control and DNA repair.
- I.e. it performs these rolls through the modulation of transcription and chromatin remodeling.
- BRCA 1 is a large protein of 1863 AA in size.
- BRCA 1 is a ring finger e3 ubiquitin ligase that has been shown to modulate DNA repair through the epigenetic chromatin remodeling and the repositioning of DNA damage response proteins on chromatin.
- BRCA1 functions through interactions with other proteins.
- In particular BRCA 1 interacts with its partner BARD 1 to modulate ubiquitylation of proteins.
- How ubiquitylation works in the control of these biological properties is not known but it is known that BRCA 1 e3 ligase can control G2/M cell cycle checkpoint control through ubiquitin dependent proteolysis.
- BRCA1 forms a lot of functional complexes with different proteins.
- It can form complexes with BARD1 to regulate ubiquitylation
- It associates with CMIC and p53 to regulate transcription.
- It can associate with ATM and ATR kinases and chk2 and 1 to regulate cell cycle checkpoint control
- BRCA2 and RAD 52 to regulate DNA repair.
- All of these interactions contribute to maintaining genomic stability at the cellular level.
68
what allow BRCA1 to engage in cell cycle checkpoints
- BRCA1 is subjectto both atm, atr and chk2 dependent phosphylation events.
- These allow BRCA 1 to engage in cell cycle checkpoint control and ds break repair.
- Phosphylation of BRCA 1 on serine 1387 by atm allows brca 1 to promote cell cyle arrest during S phase.
- Wheras atr dependent phosprylation on serine 1423 allows BRCA 1 to initiate cell cycle arrest in G2-M stages of the cell cycle.
- In is belived brca 1 can also engage with the p53 pathways to regulate G1-S checkpoint control.
- DS break reapir: the chk2 dependent phosphylation of braca 1 on serine 988 allows BRCA 1 to facilitate homogous recombination through gene conversion and inhibit the more error prone NHEJ pathway.
- These are the principle functions of brca1 in maintenance of genomic stability – regulation of cell cycle arrest ability to promote ds break reapir.
69
familial breast cancer susceptibility genes
- **BRCA2/(aka. Fanconi anaemia complementation group D1) (FANCD1)**
- BRACA2 germline mutation Predisposes to early onset breast carcinoma and ovarian tumours
- Greater incidence of breast v ovarian cancer
- Increased risk of prostate cancer, gall bladder and bile duct cancer, stomach cancer and malignant melanoma associated with BRCA2 mutations
- BRCA2 mutation 6174delT found in Ashkenazic women with BC
- **BRCA1 and BRCA2 compared**
- BRCA1 and BRCA2 make an equal contribution to early onset BC
- Somatic mutations in either gene are not a major feature of somatic breast cancer but do contribute (90% of all BC)
- With regard to the actual tumours there are differences in histology e.g. BRCA1 associated tumours have a higher mitotic rate and more lymphocyte infiltration whereas BRCA2 have higher tubule formation
70
role for BRCA2 in DNA recombination repair
- BRCA2 is a large protein of 3419 AA.
- It contains 8 internal BRC repats of approx. 35 AA in length that are required to bind the RAD 51 recombinase which functions in homologous recombination – this underlines the function of BRCA 2 in DNA recombination repair.
- BRCA 2 also contains C terminal OB folds that bind ss DNA in the dss1 protein.
- These domains allow BRCA 2 to associate with DNA.
- How does BRCA 2 function in recombination repair?
- Upon DNA damage and the formation of ds break that is then resected to form a single stand.
- The single strand is coated with RPA complex
- BRCA 2 associated with rad51 and serves to displace the RPA complex from resected ss DNA and to facilitate rad 51 association with ss DNA in an ATP-dependent manner to form rad 51 nuclear protein filaments which are essential for the homology search and to form a joint DNA molecule that comprises a processed DNA break and the repair template in a process to ensure the fidelity of the repair process which can either result in non-crossover events or crossover events but essentially these DNA molecules are the same.
71
what does BRCA1 and BRCA2 inactivation promote
- BRCA 1 is involved in cell cycle checkpoint control, ds break repair
- BRCA2 is involved in ds break repair
- It is therefore not surprising that the loss of these proteins promotes cellular radiosensitivity so cells are less resistant to radiotoxicity as cells cannot use BRCA proteins to initiate checkpoint control or DNA repair.
- This is illustrated in this figure where BRCA 1 and BRCA 2 levels in cell by RNA interference.
- The cells lacking BRCA 1 or 2 undergo cell death at lower doses of radiation
- i.e. BRCA1 and BRCA2 inactivation promotes radiosensitivity
72
what are the functions of BRCA1 and BRCA2
- BRCA1 and 2 are both involved in the DNA damage response
- Loss of BRCA1 or 2 both result in increased sensitivity of cells to ionizing radiation
- The major role of BRCA2 is to modulate DNA DSB repair through its interaction with Rad51 in recombination repair
- BRCA1 also has a role on recombination repair through interaction with BRCA 2 but additionally plays a wider role as a cell cycle checkpoint protein
