TOPIC D: Cells Flashcards
Outline Cdk signalling
Cyclin dependent kinases control the cell cycle
Cdks are activated by binding to a cyclin member
Binding moves the T-loop exposing the ATP for phosphorylation
Cyclin is broken down by Ub-proteasome which then stops Cdk activation
Outline Ub-proteasome signalling
Ub-proteasome mediated degradation.
Ub activating enzyme (E1) conjugates to Ub. Ub swaps onto E2. E3 transfers Ub onto another protein to activate it.
Poly-Ub signals for proteasome to degrade protein by using ATP
Explain Cdk role in nuclear envelope degradation in mitosos
Cdk phosphorylates the nuclear lamina subunits, changing their conformation, breaking envelope apart.
Allows cystolic condensing protein to contact DNA and condense chromatin
Explain Cdk role in microtubules in mitosis
Cdk reorganises the microtubule network from interphase into spindles.
It phosphorylates microtubules associated proteins (MAPs) to inactivate them and allow spindles to form and motor proteins to move centrosomes to the poles
Explain Cdk role in anaphase of mitosis
M-Cdk activates anaphase promoting complex (APC) which has Ub ligase activity (E3).
APC targets securin, initially bound to active separase.
Signals Ub-P degradation and allows active separase to cause separation of chromosome arms.
APC then ubiquinates M-Cdk for degradation of the cyclin. Allows phosphatases to dephosphorylate protein in cytokinesis (allows forming of nuclear envelope, interphase, microtubules). This ends mitosis.
Explain remodelling of microtubules
heterodimer of alpha and beta subunit form tubulin.
Tubulins form protofilaments which form MTs
Dynamic instability is where tubulin binds to GTP cap causing MT elongation. GTP can undergo hydrolysis which will deplete the GTP cap and cause shrinkage.
Cdk can phosphorylate MAPs which causes them to bind to the side and stop dynamic instability (stabilises MT)
Explain MTs at the centrosome
Centrosomes are where MTs radiate out. MT organising centre (MTOC) is where negative end of MT binds to gamma tubulin bound to ring complex and positive end grows to the cell periphery
Kinesin and Dynein directions of movement
Kinesins: + end directed (into cell)
Dyneins: - end directed (to the edge of cell)
What are the 3 types of MTs
Astral: anchor and position centrosome
Kinetochore: Contact kinetochore of chromosome by dynamic instability until contact
Polar/interpolar: Link to other side polar/interpolar MTs to centralise mitotic apparatus
Explain what happens at the spindle assembly cell cycle checkpoint
Mad2 sits at free kinetochores and blocks APC action
Dissociates when MT reaches kinetochore by travelling with dynein to other end.
When all Mad2 gone, APC can work
What are the 3 ways protein can be transported around the cell
gated transport by channels
Transmembrane transport through proteins
Vesicular transport
Explain nuclear pore complexes
Cystolic fibrils recognise protein
Basket on other side acts as sieve for only less than 50 kDa proteins to diffuse into nucleus
Explain the nuclear localisation signal
Basic stretch of amino acids that signals for nuclear import
Importin binds to protein and then to Ran protein inside of nucleus. Then rhe protein is released and Importin-Ran complex dissociates
How can the nuclear localisation signal be regulated
Protein binding: TF-BP binds to protein at importin site, not allowing transport
Modification of/around the NLS: cannot be recognised
What are TOM and TIM
translocase of the outer membrane
translocase of the inner membrane
transmembrane transporters of the mitochondria
Explain protein transport into the mitochondria
Signal sequence is a basic amphipathic alpha helix at N terminus.
Signal sequence binds to import receptors and travels through TOM and TIM
Signal sequence cleaved off and folding occurs once in matrix
Explain transport into the ER
Signal sequence is a stretch of hydrophobic aa followed by basic aa.
signal recognition peptide (SRP) binds to signal and brings the tRNA complex to the protein translocator by binding to the SRP receptor
Then translation feeds into the ER
How do proteins insert into the ER membrane
ER translocon opens and laterally releases hydrophobic transmembrane anchor
As protein leaves, if it has a stop sequence in it, it will dock to the membrane
What does Calnexin do
Holds onto the glycosylation of a protein in the ER membrane to ensure the protein is folded correctly and then released
Explain the deficiency resulting in cystic fibrosis
Deficiency in cystic fibrosis transmembrane conductance regulator (CFTR) which is a chloride channel.
F508del mutation causes defect that reduces protein levels due to incorrect folding in ER.
Explain treatment of cystic fibrosis
Corrector or potentiator drug treatment
ORKAMBI treament includes
(corrector) lumacaftor: chaperon
(potentiator) Ivacaftor: increases open probability of CFTR channels
How does clathrin work
Three legs make up triskelion which form around membrane into spherical shape
pulls membrane and creates vesicle
dynamin pinches the thin neck causing budding
Explain soluble ER protein retrieval
All these proteins have same sequence at C terminus
KDEL receptor binds as it leaves the ER and then forms vesicle from the golgi back to the ER
Lower pH out of ER gives receptor high affinity for protein as opposed to when in the ER
What are the 3 types of secretion at the trans golgi
Signal mediated diversion to lysosomes
Constitutive secretory pathway (usual secretion out of the cell)
Signal mediated diversion to secretory vesicles (for storage and regulated secretion)
Explain protein transport to lysosomes
Lysosomal enzymes are glycosylated in the ER
In the golgi phosphate is added to the mannose to produce M6P. This is by GlcNAc phosphotransferase.
M6P receptors recognise protein and a vesicle transports it to the lysosome
What is I cell disease
Lysosomal hydrolases are not targeted to the lysosomes and are secreted.
Cell fills up with inclusion bodies
Results in short stature, and developmental delay
What is necrosis
Accidental cell death where contents flows out of cell
Necrotic cells trigger inflammation by cells of innate immune system due to dangers of contents in EC space
This is known as danger associated molecular patterns.
What is apoptosis
Purposeful cell death from within
Cell shrinkage
Nuclear blebbing (fragmentation of nucleus)
Blebbing of plasma membrane
What are the 2 types of signals that trigger apoptosis
Inside the cell: could be to protect body
Outside the cell: other cells signal for cell to die
What are caspases
Proteins that orchestrate and execute apoptosis
Initiator caspases: activated by dimerisation and then activate effector caspases
Effector caspases: activated by cleavage and activate cell death by other proteins
What are the 2 ways of caspase activation
Extrinsic death receptor pathway
Intrinsic mitochondrial pathway
Explain the caspase extrinsic activation pathway
Death receptors undergoes conformation change when EC ligand binds causing trimerisation of 3 death receptors, recruits adaptor proteins.
FADD binds intracellularly and recruits caspase 8, forming death inducing signalling complex (DISC). Attached caspases dimerise which then activates effector caspases.
Explain the caspase intrinsic activation pathway
Apoptosome causes dimerisation and activation of caspase.
Apoptosome becomes activated by Apaf1 binding to cyt c (released from mitochondria) and these subunits coming together to form apoptosome.
Explain cyt c release regulation
Bcl-2 acts as guardian and does not allow release of cyt c from mitochondria (thus no apoptosis)
Bax/Bak when active form oligomers to produce membrane pores and cyt c release. Bcl-2 binds to Bak to prevent this.
BH3-only proteins bind to Bcl-2 allowing Bak to work
Explain common BH3 mimetics
ABT-737 binds to Bcl-2 like BH3.
Navitoclax ( a derivative of ABT-737) does this but reduces platelet count by binding to Bcl-xl on platelets inhibiting cell survival
Venetoclax binds does not bind to Bcl-xl as much but still binds to Bcl-2. Therefore it targets cancerous cells but not platelets.
