Cell Cycle 3 – Regulation of Cycle and Signalling
Anil Chopra
1. Explain in molecular terms the concept of a tumour suppressor, using as an
example the retinoblastoma (Rb) susceptibility gene product.
2. Explain in molecular terms the concept of a proto-oncogene and provide
examples.
3. Explain how apoptosis can limit inappropriate cell division and discuss the role of
cytoprotective pathways (e.g. the phosphoinositide 3' kinase pathway) in
facilitating cell proliferation.
 The turnover of haematopoietic
cells in a 75kg adult is around
500 000 000 000 (500 billion)
cells a day.
 A small number of progenitor
stem cells (1% of marrow cells)
maintains production.
 The stem pool responds to
external stresses e.g. in
hypoxia, number of red blood
cells increases. In bacterial
infection granulopoiesis
increases.
With the use of proprium iodide staining, we know that the most DNA is in the cell
when it is in G1.
E2F is a transcription factor that regulates genes. When the DP1/E2F1 heterodimer
is bound to pRB (a tumour suppressor), it cannot initiate transcription, however, when
certain cdks (cyclin dependent kinases) bind with certain cyclins, then they
phosphorylate the pRB causing the DP1/E2F1 to detach from the pRB leaving the
DP1/E2F1 to initiate transcription. In this way pRB acts as a tumour suppressor.
Regulation of DP1/E2F1 heterodimer during the cell-cycle
pRB
G1
DP1
E2F1
Transcriptional inactive/
DNA bound
cyclinD/cdk4 or 6
cyclinE/cdk2
cyclinA/cdk2
pRB
DP1
G1/ S
Transcriptional active/
DNA bound
E2F1
cyclinA/cdk
2
DP1
E2F1
S/
G2
Transcriptional
not DNA
inactive/
bound
In order to control the cell cycle we need to use cyclin-dependent kinase inhibitors
(CKI’s) of which there are 2 types:
 INK family – cause separeation of the
CDK from the cyclins.
 Cip/Kip family - bind to and inhibit the
action of the CDK-cyclin complex.
M
G1
Cyclin BCDC2
P
pRb
E2F
G2
P
Ink4b
X
P
P
Cyclin D-CDK4/6
pRb
E2F
S
p21Waf-1
p27Kip-1
p57Kip-2
Cyclin E-CDK2
p21Waf-1
p27Kip-1
p57Kip-2
Cyclin A-CDK2
At restriction point “R” in the cell cycle, the presence of growth factors is needed for
G1 to progress. In the presence of growth factors, cyclin D binds to CDK 4 and
phosphorylates the pRB (tumour suppressor). Later in the mid/late G1 phase, the
complex formed when cyclin E binds to CDK 2 causes the pRB to be phosphorylated
again resulting in its release of the DP1-E2F1 complex.
Early G1
CDK4
pRB
G1/S
Mid/Late G1
Cyclin D
P
CDK2
pRB
Cyclin E
P
pRB
P
P
P
DP1
DP1
E2F1
D
DP1
P
E2F1
E2F1
Growth factor
Cyclin E
Cyclin E
The binding of CDK2 to cyclin E also causes
the breakdown of the CDKI p27.
All E2F genes
Mid/Late G1
CDK2
Cyclin E
P
p27
p27
An oncogene is a mutant gene which promotes cell proliferation. A proto-oncogene is
the normal cellular gene corresponding to the oncogene.
There are a number of proto-oncogenes that are involved in normal cell division, that
once mutated become oncogenes and allow uncontrolled proliferation of cells (i.e.
cancer). These include:
 The EGF receptor
 Ras which may become the oncogene V12Ras following a Gly12Val mutation, or
L61Ras following Gln61Leu mutation.
 C-Raf which becomes the oncogene v-Raf upon deletion of it regulatory domain.
 C-Jun which becomes the oncogene v-Jun upon deletion of its regulatory domain.
Mutations usually result in cell cycle arrest and apoptosis. P53 is a tumour suppressor
gene, and it also promotes apoptosis. For unregulated cell signals to occur, one of the
following must occur:
o There must be an active proliferative pathway
o Cell cycle arrest must be inhibited
o Cell survival signals must be activated.
The ERK cascade is cytoprotective as it produces a protein called p90RSK which can
phosphorylate Bad. This means that Bad is bound to the protein and can therefore not
be part of apoptosis. This leads to the proliferation of the cell.
PI3K (phosphatidylinositol 3-kinase) promotes cell survival. A extracellular survival
signal activates a receptor tyrosine kinase, which recruits and activates PI3K. It results
in the phosphorylation of the Bad protein, which holds inhibitory proteins. These are
released and can block apoptosis, thereby promote cell survival.
The diagram shows the activation and
inactivation of Ras. However, V12Ras
prevents GAP binding to it, so once the
Ras has been switched on, it cannot be
switched off, therefore Ras will
continuously stimulate the ERK cascade
which could lead to uncontrolled cell
proliferation.
A similar thing occurs with L61Ras – it
prevents GTP hydrolysis (i.e. stops the
GAP from doing its job) and it to will
therefore remain active continuously.