CBG Lecture 9: Cell Cycle Flashcards
what is the cell cycle
a four phase internal clock in the cell that regulates cell division
which domain is it found in
eukaryotes
what are the four phases of the cell cycle
INTERPHASE: G1 - growth S: Synthesis G2: growth and prep for division MITOSIS/MEIOSIS + CYTOKINESIS
what stages does interphase include
growth G1
synthesis
growth and prep for division G2
how long does G1 phase last
8 hours
how long does synthesis phase last?
DNA replication lasts c.12 hours
what happens ni synthesis phase
dna replication
eg. diploid to tetraploid
which phase is the longest in the cell cycle
synthesis: c.12 hours
what happens in G2 phase
growth and prep for cell division c. four hours
what happens in M phase
meiosis/mitosis/cytokinesis
how long does M phase last
half hour
what evidence exists for M phase
its histologically obvious - chromosomes condense
can see stages of meiosis and mitosis
what evidence is there for S phase
identify by 3H- thymidine autoradiography
50% of cells normally labelled, shows cells spend 50^ of their time in S phase
how can G1 and G2 phases be identified
use flourescent dyes to measure [DNA]
use flow cytometry
what is flow cytometry
each drip from vibrating nosel adds just a single cell, so if you add a flourescent dye, can tell which phase youre in by amount of flourescence
what is flow cytometry used for
measures [DNA] can see which phase of cycle a cell is in
which colour flourescence do cells in G2 phase show in flow cytometry
green
compare the amount of flourescence in G1 cells to G2
G2 has double amount of DNA than G1 therefore will have double the flourescence
which cells are used for fusion experiments
HeLa cells
what are HeLa cells
cervical cancer cells - immortal cells
what was the main result from fusion experiments
S phase nuclei induce S phase in G1 but not in G2
what is the result of a fusion experiment with a cell in S phase and a cell in G1 phase?
S contains factors that drive G1 to replicate DNA so get 2 S phase cells
what is the result of a fusion experiment with a cell in S phase and a cell in G2 phase?
G2 are refractory to the S phase factors, so no change to cells
what is the result of a fusion experiment with a cell in G2 phase and a cell in G1 phase?
no change
what did injection studies in frog oocytes show
maturation promoting factor can induce M phase in G2 nuclei
what does maturation promoting factor do
induce M phase in G2 nuclei
if you have a G2 cell and place in external control progesterone into it, what happens
induces M phase
how can M phase be induced
by adding progesterone
what does microinjection of M phase cytoplasm into G2 cells do
induces M phase
what can cell division cycle mutants do
help to understand cell cycle regulation
what are checkpoints in a cell cycle
restricition point in mammalian cell/start point in yeast where Cdc21/ Cdk1 all work
what happens to the cell cycle of yeast mutants
temperature sensitive yeast mutants arrect in G1 at high T
what is human homologue of Cdc28
Cdk1
what phase do temperature sensitive yeast mutants go to at high temperatures
G1 go to G0 at high temperatures
what are CDC mutants? what does CDC stand for
cell division cycle mutants Cdc28
what does Cdk stand for
what can an arrest to Cdk1 do
cyclin dependent kinase
Cdk1 can cause arrest to G1 phase
what are check points in a cell cycle regulated by
internal and external factors
how many check points in the cell cycle are there
what are they
3
G1
G2/M
Metaphase
what internal/external growth factors taken into account at G1 checkpoint
internal growth factors: cell size
external growth factors: growth hormones, hormone factors, check whether DNA is damaged
what internal/external growth factors taken into account at G2/M checkpoint
internal: check all DNA present (can arrest at this point+wait/grow DNA)
is all DNA replicated
cell size
dna damage
what internal/external growth factors taken into account at metaphase checkpoint
chromosome alignment
how do you rectify misaligned chromosomes in metaphase checkpoint
use tubulin cytoskeleton
what are cyclins
proteins that cycle in concentration over the cell cycle
give an example of an S-type cyclin
cyclin D - which cycles in sea urchin embryos
how does [S-type cyclin] change over time
levels build up from G1 through to S through to G2 and drop dramatically at G2/M checkpoint
what is progress through the cell cycle controlled by
a cyclin dependent kinase
whatt do cyclins do
detrermine the specificity of the CDK for its protein targets
how many CDKs do yeast have, what?
yeast have a single CDK, vertebrates have several
what can misregulation of CDKs result in
uncontrolled growth - cancer
what are CDKs given specificity by
cyclins
why wont CDKs work alone
cyclins tell CDKs what they should be modifying and which proteins they should switch on/off
enzymes to activate etc
what is the passage through metaphase checkpoint associated with
the destruction of M cyclin by APC: anaphase promoting complex
what is APC - what does it do
anaphase promoting complex
associated with the destruction of M cyclin
what complex promotes passage through start/G1 checkpoints
CDK+ G1 cyclin
and CDK+G1/S cyclin
how does first complex promote passage through start/G1 checkpoints
CDK+ G1 cyclin
and CDK+G1/S cyclin
oassage through start by phosphorylating
eg. Retinoblastoma protein which frees EF2 - transcription factor
give an example of passage through start checkpoint of cell cycle
CDK + G1/S cyclin
retinoblastoma protein which frees EF2
how do CDKs become activated
association with a cyclin makes it partly active
phosphorylation by CAK makes it fully active
how does a CDK become partial active
association with cyclin
how does a CDK become fully active
after association with cyclin, becomes phosphorylated by Cak
how can CDK be both inactivated AND activated by phosphorylation
has multiple phosphorylation binding sites
how is CDK inactivated
phosphorylation of 2 phosphate groups by Wee1
How does CDK+S cyclin complex initiate synthesis
phosphorylation
give an example of CDK+S cyclin complex initiating synthesis
histone mRNA stem-loop binding protein-stabilises histone mRNA
how is DNA synthesis initiated
by CDK+S cyclin being phosphorulated
what promotes passage through G2/M checkpoint
by CDK+M cyclin (maturation promoting factor)
what is MPF made of
maturation promoting factor
CDK+M cyclin
how is passage through G2/M checkpoint promoted
MPF (CDK+MCyclin) phosphrylation
eg. nuclear lamins - break down nucleus
myosin - prevent cytokinesis
condensin = condenses chromosomes
what happens when cyclin synthesis>degradation
cyclin accumulates and can force cyclin CDK complex to go throughcheckpoint
how is a CDK/cyclin complex broken down
cyclin ubiquitination by APC -anaphase promoting complex - and degraded
cyclin degraded, CDK is recycled
how are cyclins regulated
by a balance of synthesis and degradation
what pathway is used to regulate cyclins
MAPKinase/K/K patheay which results in G1 cyclin expression
which pathway results in G1 cyclin expression
MAPK/K/K Growth factor cascade
what are CDK/cyclin complexes inhibited by
Cdk inhibitor proteins (CKIs) like p21
what is a CKI
Cdk inhibitor protein
name a CKI
p21
what is p21
a CDK inhibitor protein
how can a mutation be detected
p53 presence
what can a mutation in p53 and p21 do
more likely to replicate damaged/mutated DNA -> lead to cancer
name some tumour suppressor genes
p21
p53
what is a tumour suppressor gene
gene that encode proteins that act as brakes on cell cycle
what type of mutation does a tumour supressor gene need to become dangerous
recessive mutation - both copies need to be broken
give ecamples of tumour supressor genes
genes encoding p53
retinoblastoma protein
what is an oncogene
gene that if a dominant mutation exists, it will accelerate cell cycle progression
give ecample of oncogene
genes encoding Ras, RTKs,Myc
how is an oncogene mutation dangerous
it generates the genes to be constituently on regardless of promoters/operatiors/repressors
