Ch. 8: RB and Cell Cycle Flashcards
Cell cycle
map of all the steps
that occur within a cell from the moment it is created from one cell division to the end of the next round of cell division
Contact dependent growth
If you put human cells in culture they will continuously divide until they run into each other
Mitosis
Process of cell division that includes 1. equal division
of chromosomes between 2 daughter cells 2. the physical
splitting of 1 cell into 2 daughter cells. (~1 hour)
Stages of mitosis
- prophase
- prometaphase
- metaphase
- anaphase
- telophase
Cell growth and example
cells are getting physically bigger in preparation for cell division
- Ex: Doubling macromolecules and organelles- (lipids- membrane, proteins- ribosomes, carbs- modifications on cell proteins), organelles-endoplasmic reticulum
Synthesis
Cells are duplicating their
DNA in preparation of cell division (DNA polymerase) (varies)
Stages of cell cycle
- mitosis
- G0
- G1
- synthesis
- G2
G0 (Temporary and permanent)
Cells make decision not to divide at that moment in time. (Can occur during
G1 as well.)
* temporary- quiescent occurs at G0
* permanent- senescent occurs at G1/2
Gap 1
Gap between G0 and Synthesis (~6-8 hours)
Gap 2
Gap between Synthesis and Mitosis
(~3-5 hours)
Interphase
G0, G1, S, and G2
(part of the cell cycle not containing mitosis)
What would happen if chromosomes are not duplicated properly?
- potential transformation
What would happen if division does not occur properly and each daughter cell does not get exactly 2 copies of every chromosome?
- If 2+ copies, proto-oncogene can become an oncogene
- if 2- copies, tumor suppressor genes could cause transformation
In a way, proteins/molecular complexes involved in the cell cycle are tumor
suppressors because…
loss of them would
lead to situations that promote cancer
Very important for the cell to have # of quality control steps along the cell cycle to
ensure…
cells are ready to go on to the next step. (Alleviates potential for cancer).
checkpoints
control mechanism to ensure the next step in a process does not proceed
until necessary prerequisites have been fulfilled.
R point
The most critical decision-making occurs whether
to continue dividing or exit the cell cycle (and
enter G0). This occurs during G1 up to the R point.
Cells are listening to their neighbors
Example of issue at a check point
- DNA damage G1 to 2
- DNA damage S to G2
- entrance to M is blocked if replication is not completed
- anaphase is blocked if not attached to the mitotic spindle
the most critical checkpoint regarding the onset of cancer (and example)
the R point
- Ex- Anchorage dependent growth: ensure cells are properly attached to ECM (via integrins) before S can occur. If cells are not attached at all initiates self-destruction (apoptosis)
- Anchorage independent growth- property of transformed cells- involves oncogenes (such as RAS and SRC) tricking the cell into thinking it is properly anchored
How do cell cycle checkpoints work on a
molecular level?
- CDKs?
Cyclin dependent Kinases
- CDK’s are usually present throughout the cell cycle.
- Each CDK phosphorylates and activates a set of target genes involved in a specific
stage of the cell cycle
What regulates CDK activity?
the presence of cyclins
cyclins
proteins that bind to CDK’s and activate them (turn them on)
1. Activate their kinase activity
2. Help them find the right substrates to phosphorylate
When are cyclins present/active?
at precise times in the cell cycle
Ex- CyclinB is only present at the M stage, therefore CDK1 is only active during this stage
Which CDK and cyclin do you predict to be important
during the R check point?
CDK4/6
What regulates cyclin D1 heavily?
extracellular inputs
(Wnt, cytokine, hedgehog, other ligands)
What does growth factor RTK do to D1?
RTK activation ultimately increases the transcription of D1
What does cell attachment to the ECM do to cyclin D1?
Cyclin D1 txn is also regulated by cell
attachment to the ECM (anchorage dependence growth)
Why does D1 have to be heavily regulated?
D1 needs to be heavily
regulated because once CDK4/6 is active, cell division WILL occur
Cyclin-CDK complexes are regulated by…
inhibitors
Different terminology over the years
* Originally- INKs (inhibitors of CDK4)
* Currently- CKI (cyclin kinase inhibitors
Function of CKI
inhibit Cyclin-CDK formation
and function
P21CIP
cyclin dependent kinase
inhibitor
- Levels of p21CIP increase in a cell in response to cellular stresses including DNA damage
- Will shut down E-CDK2/A-CDK2 complexes so that the cell cycle will stop until DNA is repaired
What inhibits CDKIs
growth promoting signals
- Mitogens (such as growth factors)- activate RTK’s and subsequent activation of AKT
- Active AKT phosphorylates p21Cip1 and p27Kip1 in the nucleus which causes them to be exported to the
cytoplasm - Lack of CDK inhibitors in the nucleus causes subsequent activation of
cyclin-CDK complexes in the nucleus - Promotes progression of the cell cycle (cell division)
When in the cell cycles is Db de-phosphorylated?
during G0
When in the cell cycles is Db mono-phosphorylated?
during the G1 stage of the cell cycle up until the R checkpoint
When in the cell cycles is Db hyper-phosphorylated?
after the R checkpoint and at all other stages of the cell cycle
What is RB important for?
for entry past the R checkpoint
Viral E7 protein of HPV binds to RB and inhibits…
its function and targets it for proteasomal degradation
RB and TP53 usually inhibit…
cell proliferation and
ensures cell division is
tightly regulated (it only
occurs when necessary)
Viral proteins bind to RB
and TP53 proteins, this
allows for…
for the transcription
of genes that promote cell
division (proliferation)
What inhibits E2F TF’s? how?
De-phosphorylated and mono-phosphorylated
(active) RB
Active RB (de or
monophosphorylated) binds to E2F pocket proteins and recruits HDAC which changes the chromatin in such a way that inhibits RNA polymerase (txn)
what are E2F pocket proteins?
are transcription
factors (TFs) that regulate genes that cause transition from G1 to S phase
What prevents RB from binding to E2F?
hyper-phosphorylation
What happens if RB cant bind to E2F?
- Thus histone acetylases can bind to E2F
- This changes the chromatin in such a way that now RNA polymerase can bind
these genes and txn is activated - Reminder- these genes cause transition from G1 to S phase (transition past the R
point) (~500 genes)
What controls RB phosphorylation?
- de-phosphorylation: protein phosphatases
- mono-phosphorylation: CDK4/6 and cyclin D (active G0 CDK’s)
- hyper-phosphorylation: CDK2 and cyclin E (active G0 to S1 CDK)
Determine which version of RB is present in each part
of the diagram below labelled in purple
(slide 32)
1- de-phosphorylated
2- mono-phosphorylated
3- hyper-phosphorylated
4- hyper-phosphorylated
5- hyper-phosphorylated
6- hyper-phosphorylated
How does GF-RTK signaling induce cell division?
Growth factors -> RTK receptors -> Ras -> Cyclin D1 and E -> inactivation of RB -> activation of E2F’s -> S-phase entry
RB pathway Review
- Un-phosphorylated RB puts cells in G0 (out of the cell cycle)
- RB is monophosphorylated by active CyclinD-CDK4/6 during the progression of G1
- RB is hyperphosphorylated by active CyclinE—CDK2 during the end of G1 and
beginning of S (R checkpoint) - Once RB is hyperphosphorylated, it no
longer binds E2F TF’s - Active E2F activates the txn of genes involved in S phase, ex- DNA polymerase
What does E6 and E7 inactivate?
TP53 and RB (tumor suppressors)
What does E6 and E7 do in HPV?
they integrate into the host genome and are expressed at high levels
MYC transcription factor from previous lectures
- Tumor virus lecture-
-> B-cell lymphomas are induced by ALV integration in front of the MYC locus in
the host genome (slow transforming retrovirus)
-> Increases the gene expression of MYC - Oncogene lecture-
-> Burkitts lymphoma a chromosomal translocation causes MYC to be under
the control of an active
immunoglobulin gene promoter
General MYC knowledge
- More than 70% of human tumors overexpress MYC or one of its orthologs
- Normal function- growth
promoting TF (transcription factor) - Active when heterodimerizes with MAX TF
- Growth factors cause increase in cellular levels of MYC
Deregulated MYC is
sufficient to…
drive cell proliferation in the absence of growth factors because it has many downstream effects that promote cell cycle progression
How does MYC promotes cell cycle progression
through the activation
of D2 and CDK4
- MYC/MAX promote the txn of D2 and CDK4
- Active CyclinD2-CDK4 complexes mono-
phosphorylate RB to drive progression through G1
How does MYC promotes cell cycle progression
through the activation
of Cul1
- MYC/MAX promote the txn of D2 and CDK4
- CUL1 helps degrade p27 through ubiquitylation
- p27 is normally an inhibitor of CyclinE- CDK2 complexes (which normally phosphorylate and inactivate RB)
- End Result- Inactivation of RB, further drives cells to progress through R point
How does MYC promotes cell cycle progression
through the inhibition
of p15
- MYC/MIZ-1 inhibits the txn of p15
- p15 is normally an inhibitor of active CyclinD2-CDK4 complexes
- End result- increased activity of cyclinD1-
CDK4 which mono-phosphorylate RB and
drive progression through G1
How does MYC promotes cell cycle progression
through the activation
of E2F
- MYC/MAX promote the txn of E2F transcription factors
- Further drives the progression of cells
past the R point
In summation, deregulated MYC…
Deregulated MYC is
sufficient to drive cell
proliferation in the absence
of growth factors because it
has many downstream
effects that promote cell
cycle progression
The RB pathway is dysregulated in…
most if not all cancers
About the RB pathway
- Major characteristic of cancers- uncontrolled cell growth
- Since RB control of the R checkpoint is what controls if a cell will divide or not
- Makes sense RB is dysregulated in so many cancers
- RB pathway can be dysregulated in many ways
-> (Perturbation of R-point transition in human tumors)
Perturbation of R-point transition in human tumors:
Molecules in red are hyperactive in some cancers
Molecules in blue are in activated in some cancers
Active TGFB signaling inhibits…
cell division
- Because p15 is an inhibitor of CyclinD/CDK4, activated TGFB signaling causes inhibition of G1 cell cycle
progression
