Module 5 second half Flashcards
cdc 2
cyclin dependent kinase (CDK)
Cdc 2+ = wild type = wee form
Cdc 2- = recessive = excess of wee1 or deficit of cdc25
Cdc 2D = deficit of wee1 (kinase inhibits mitosis) or excess of cdc25 (phosphatase - drives mitosis)
Kinase and phosphatase cascade
controls entry into mitosis
Wee 1 - phosphorylates Y15 of mitosis cdk, inactivating it
CAK - activated the Miro to f kinase by phosphorylation
Cdc25 - removes an inhibitory phosphate from mitotic cdk
Mitosis promoting factor (MPF)
mitotic cyclin/CDK
produced in G2 but kept phosphorylated (inhibited) until cell ready for mitosis
Checkpoints
- points in cell cycle where cell can stop if something has gone wrong
- driven by proteins that inhibit cell cycle progression
Mitotic checkpoint
DNA replication incomplete
ATR1 – CHK1 – (inactivates) – Cdc25
(ATR1 - checkpoint protein that is activated by single stranded DNA)
G2/M summary
- yeast mutants identified crucial players in G2/M transition, including activators and inhibitors of mitotic cyclin/CDK.
- phosphorylation of different CDK residues can inhibit and activate the mitotic cyclin/CDK.
- checkpoints ensure that cell has complete one ask before initiating the next. they target the major regulators of the cell cycle
Mitotic cyclin/CDKs and nuclear envelope
- nuclear lamina support nuclear envelope
- lamins A,B and C form lamina structure
- mitotic cyclin/CDKs phosphorylate the lamin proteins, causing meshwork to breakdown
APC/C ubiquitin-protein ligase
ubiquitinates mitotic proteins at key stages of mitosis
1. induces anaphase - metaphase-anaphase transition
2. induce late steps in mitosis (late anaphase) (along with Cdc14)
Phosphatase PP2A
associated with centromeres and prevents cohesins near the centromere from being removed by phosphorylation - therefore they must be removed another way
Metaphase/anaphase transition
separation of chromatids
APC/C initiates this by inducing cohesin removal at the centromere.
(ring that surrounds chromosomes)
- cohesively keeps everything together (to remember)
- cohesins are composed of smc proteins and Scc1
- separase is a protease that cleaves Scc1
- securin binds separase, inhibiting it
- when all kinetochores bind MTs, Cdc20 binfd to APC/C
- APC/C polyubiquitinates (dooms) securin
- separase, uninhibited, cleaves Scc1
- chromaids are free to separate. this define anaphase
Degradation of mitotic cyclins
APC/C specificity changes late in anaphase once chromatids have separated
Cdc1 becomes active and leads the APC/C complex to the mitotic cyclin for degradation
Ubiquitination in G1-S phase transition
CKI binds to CDK/cyclin - inactive
SCF targets CKI and ubiquitinates it so then the CDK/cyclin is now active
- ready for S phase (?)
3 mechanisms of cell specification
cell to cell interactions
- cell beside you determines your fate
asymmetric
- identical inside but ones slightly larger
morphogen gradients
- Bicoid inhibits caudal = which creates tails
- nanos inhibits hunchback
CAM
Necrosis
unplanned cell death
- rupture of cell and spilling contents to surrounding tissue
- caused by variety of attacks on cell
- toxins, burns, infection, inflammation
Apoptosis
programmed cell death
- cell chopped up and packaged for removal. DNA fractionated, cell membranes pinch off into small structures
- fragments labelled for macrophages so they cleaned up through phagocytosis
- EGL1 is produced by apoptosis signal
- EGL1 binds to CED-9, releasing CED-4
- CED-4, released from mitochondria; forms octamers in cytoplasm
- CED-4 octamers convert an inactive CED-3 precursor to active CED-3
- CED-3 is caspase, destroying various proteins
CED-3
when active:
- cleaves lamins (nuclear envelope dissolves)
- activates endonucleases (DNA is digested)
- attacks cytoskeletal components (cell structure)
- attacks cell-cell adhesion proteins (anchors for membrane)
- cleaved (activates) itself (runaway chain reaction)
CED-3 mutant
no apoptosis
CED-4 mutant
no apoptosis
CED-9 mutant
all cells die
ED1 mutant
no apoptosis
Phosphatidylserine (Ps)
marks apoptotic particles for phagocytosis
- PS is moved to outside of membrane of apoptotic cells, marking them for phagocytosis by macrophages
cancer cells select for mutations in two types of genes
proto-oncogenes
- genes that normally promote cell growth. when mutated or amplified they become oncogenes
tumour-suppressor genes
- normally inhibit cell cycle progression. when theyre mutated, cells divide out of control
Retinoblastoma
- rare childhood cancer
- heritable and sporadic cases
- caused by mutations in Rb gene
= loss of heterozygosity
The G1 restriction point is a target of cancer
- Rb rlly important for G1 of cell cycle
- 2 major kinases in G1 that promote entry into S-phase = G1 CDK + G1/S CDK
- these kinases phosphorylate Rb
(if no mitogens = go into G0) - restriction point (R or START): cell committed to another round of cell division and is no longer mitogen sensitive
- Rb represses E2F-E2F promotes expression of replication genes
- G1 phosphorylates Rb and Rb falls off E2F
Rb
poor substrate = needs to be phosphorylated multiple times
a repressor of E2F txn factor
Inactivation of APC/Cdh1
(new commitment point?)
APC/Cdh1 needs to be inactivated before cells enter S-phase
- G1/S CDK phosphorylated Cdh1
- Emi1 leads another Ub ligase to destroy Cdh1
p16
binds CDK, preventing cyclin/CDK joining. Expression of p16 is driven by stresses placed on the cell
G1cyclin/CDKs
phosphorylate (inactivat) Rb
E2F
A TF that drives expression of S-phase genes
p53
gatekeeper of cell
- detects conflicts in cell and can activate several pathways to fix them
- loss of p53 doesn’t make a cancer cell, it predisposes a cell to cancer. occurs through loss of tumour suppressor function
- ….
transcriptional regulation
- distinct genes can be turned on or off
- distinct mRNAs translated or repressed and localised to different regions of the cell
epigenetic regulation
marks on DNA that regulate gene expression and are inherited by daughter cells, but independent of DNA sequence
permanently bind Rb to E2F = cell cycle arrest
Rb pathway
- cell signals that it wants to go unto S-phase and so P16 which is inhibiting G1/S CDK goes away (depends on stress of cell - once cell is no longer stressed it released and this continues)
- g1 phase = G1/S cyclin binds to CDK and creates complex
- G1/S cyclin CDK phosphorylates Rb (which is inhibiting E2F)
- Rb leaves E2F and it is activates to create TFs for DNA replication and go into S phase
Terminal differentiation
permanent cell cycle exit
- differentiated cells become refractory to proliferate signals
- G1/CDK inhibitors and Rb family members play major roles in cell cycle exit
- terminal differentiation is distinct from senescence
(td = permanent withdrawal from cell cycle)
Cancer mutations
Gain of function = promote proliferation
Loss of function = growth suppress, avoid checkpoints and genomically unstable
Dominant negative = act like loss, mutant bind to wild type
Regulation of gene expression
Epigenetic: histone acetylation (opens chromatin) and methylation (closes chromatin)
Transcriptional: proteins that promote or inhibit gene expression
