L18&19: Cell Cycle and its Control Flashcards
Ouline the phases of the cell cycle
-G1: metabolic changes prepare the cell for division. At restriction point cell is committed to division and moves into S phase. Lasts ~10 hours
- S phase: DNA synthesis replicates genetic material. Each chromosome now consists of 2 sister chromatids. Lasts ~6 hrs
- G2 phase: metabolic changes assemble the cytoplasmic material necessary for mitosis and cytokinesis. Lasts ~3/4 hrs
Interphase now occurs (PMAT)
- M phases: mitosis (nuclear divsion) followed by a cell division (cytokinesis). Lasts ~2 hrs
How does the cell cycle process work?
- process is unidirectional, only other point is cell death
- driven by series of cell cycle kinases (CDKs)
- regulated by checkpoints to prevent erros
- spindle assmebly checkpoint occurs at end of M phase
- restriction point is just before G1
Explain the strucuture of chromosomes
- centrosome is a constricted region of chromosme containing specific DNA sequence, to whivh is bound 2 discs called kinetochores
- kinetochores serve as points of attachment for microtubules that move the chromosomes during cell division
Explain the use of sea urchins in cell cycle research
Sea urchin eggs were used in the first demonsration of periodic protein degradation, a concept whihc is fundamental to the cell cycle
Explain the use of yeast in cell cycle research
Yeast via genetic mutation analysis has been used to identify nearly all cell cycle regulatory genes
Explain the use of xenoupus eggs in cell cycle research
Xenopus eggs and embryos extracts have been used to make biochemical discoveries
- one discovery being of mutation promotion factor (MPF), now commonly referred to as CDK1 activity and is the key protein kinase that controls cell cycle
Explain the use of drosphila in cell cycle research
Drosphila embryos can be used for knockin/out genes of interset and mammalian cells in culture
What drives the cell cycle?
The rise and fall of several protein kinase activities
- cdk activity is dependent on 4 hroups of cell cycle specific cyclin:
- cyclin D1,2,3 (cdk4&6): G1
- cyclin E (cdk2): G1/S
- cyclin A (cdk2,1): S
- cyclin B (cdk 1): M
How is the rise in cdk governed?
- rising cyclin levels (affects activation of cdk partner): cyclin txn & translation result in production of cyclin proteins which partner with relevant cdk and drive that phase of cell cycle
- dephosphorylation of cdk-cyclin complex
- specific inhibitor proteins of the cdk
What molecules usually phosphorylate residues in cdk?
Wee1 and Myt1
- to remove thesse we have cdk25 (phosphotase protein) which has 3 forms
cdc2A: CONTROLS G1/S & G2/M
cdc25B: controls G2/ M
What is the function of cks1 protein
Adaptor to target cells to phosphoproteins and mediates APC interaction with cyclin A and B
- enhances phosphorylation of cdk substrates
- has 2 forms: cksHs1 & 2
What is cdk1 activation based on?
+ve feedback: trigger activates cdc25 resulting the +ve feedback loops
- pushes cyclin kinase into active state, removing phosphorylation
- once complex is activated it further activates cdc25
Name and explain a key mechanism in cell cycle and mitsosis
Protein degradation and is acheived by ubiquitin- mediated protein destruction, which is a multist
* refer to notes for image
What is the function of SCF?
Its an E3 ubiquitin ligase which controls the G1/S transition
- formed Cu11, skp1 & Rbx1/ Roc1 subunits
What is the function of anaphase promoting complex?
Controls the metaphase- anaphase transition\
- targets are cyclin A&B plus securin
Explain how cycle control occurs in the S phase
Specifc cdk inhibitors (cdk 4&2: cyclin d&e and p21) holds cycle at G1
- in this period GF and mitogenes promote cell division and increase in concentration
- when they get to a certain level they can break this inhibition, resulting in activation cascade of 2 cdks
- this phosphorylates protein RB/ p107 releaseing key txn factor E2F which causes expression of genes needed for S phase
- synthesis of cyclin A (cdk2) and cell goes into S phase
Explain the synthesis of the leading and lagging strand
Synthesis is asymmetrical
- leading strand follows direction of unwinding at replication fork
- lagging strand has to be synthesised in opposite direction
- helicase causes unwinding and in its wake replication protein A molecules bind, at which point the primase can come along and synthesise primer for new DNA
How is the synthesis of the leading and lagging strand made more efficient?
Replication complex, including polymerase, is held in place by sliding clamp
- primer is loaded to make fragments, process extends and when everything has been replicated DNA ligase joins up okazaki fragments
Explain the assembly of prereplicative complexes
- We have origin recognition complex, Orc1-6 binding at origin
- Then cdt1 and cdc6 join and then so does helicase
cdt1 and cdc6 joining is mediated by germinin, which binds to cdH preveting it from causing the bnding of helicase so unwinding can no longer occur
- germinin accumulates in S phase
How does telomerase synthesise DNA at chromosome ends?
- they add several repeat DNA regions on lagging strand to serve as priming region, so lagging strand can be completed
- resulting strucuture at end of each chromosome known as telomere
- shortening of telomere is associated with aging
How are nucleosomes packaged in the final stage of S phase?
- nucelosomes have ~147bp of DNA wrapped around each octamer to make a nucleosome
- then tightly packed into a 30nm fibre and further packaged by contraction
Histones have protuding N-terminal tails. True or false?
True, these are modified post translation and involved iwth gene expression
- can also involve non-histone proteins (and acetylation to modify chromatin structure)
Explain the process of nucleosome assembly
- Acetylated H3-H4 tetramer complexes with assembly factor CAF-1
- This is recruited to replication fork and incorprated into new DNA
- H2A& B dimers loaded on the H3-H4 tetramer via another assembly factor NAP-1 to form octomer
Name particular histone modifications associated with differnt functions
- histone acetylation: active genes
- H3K4me3: active promoters
- H3K9me3 and H3K27me3: heterochromatin
- H3T3ph and H3S10ph: mitosis
- H2AX.S13ph (gamma H2AX): DNA damage
What can be found on histone tails?
Markers which can have
- writers: enzymes which coe along and deposit phosphorylation
- erasers: gets rid of phosphorylation thus marker
- reader: recognises specific phosphorylation and bind
