Protein Trafficking Flashcards
What can polarity be maintained by?
- physical barrier (AIS)
- selective trafficking
- selective retention
- selective degradation
- local translation
What is localised translation?
when certain proteins are synthesised away from the cell body
When does the axon initial segment (AIS) form?
after axon establishment
What does the molecular sieve do?
- prevent free diffusion of selected membrane and cytoplasmic proteins into axons
- regulate the initiation of APs
What does the AIS have a similar molecular organisation to?
node of Ranvier
What is the key difference between the axon hillock and the AIS?
the axon hillock is present in the cell body of the neuron while the AIS is present in the proximal part of the axon
What is the role of the AIS in polarity maintenance?
- serves as a submembrane diffusion barrier that restricts the mobility of plasma membrane components
- acts as an intracellular selective filter for the transport of organelles and molecules through the cytoplasm
What is inter-compartmental transport essential for?
communication and support
Why are proteins and organelles in constant motion?
- synapse formation (from sites of synthesis to sites of function)
- for degradation
- for everyday requirements
In axonal transport, where is the plus and minus end respectively?
- minus = cell body
- plus = extremities
What are kinesins?
plus end motors with 45 genes that take newly synthesised proteins from the cell out to the axon terminal
What is kinesin-13 and 14 respectively?
- 13 = MT depolymerising (catastrophe promoting)
- 14 = minus-end directed
What are the 3 major domains of a kinesin motor?
- tail (cargo binding and auto inhibition mediation)
- stalk (dimerisation domain)
- head (motor domain)
What are the 6 steps of kinesin walking along an MT?
- in solution, both kinesin heads contain tightly bound ADP and move randomly
- when one of the heads (M2) encounters an MT, it binds tightly and releases ADP
- ATP rapidly enters the nucleotide binding site which triggers the neck linker to zipper onto the catalytic core
- this action throws the second head (M1) forward and brings it into contact with the next MT binding site
- the trailing head hydrolyses ATP, releasing phosphate
- the neck linker unzips from the trailing head and the leading head exchanges its nucleotide which allows it to zip to the catalytic core and the cycle repeats
How does the tail of kinesin bind the cargo?
via adapter proteins
Give an example of a cargo that binds directly to the tail domain without the need of adapter proteins
SNAP25
How can kinesin autoinhibition be relieved?
by cargo binding and phosphorylation mechanisms
How do autoinhibited kinesins work?
dual inhibitory mechanisms control kinesin-1, 2 and 3 motors through domains that inhibit MT binding and domains that inhibit processive motility
How can kinesin-1 motors be reactivated after autoinhibition by cargo binding?
the interaction with FEZ1 and JIP1 that releases the restraints of MT binding and processive motility
How can kinesin-7 motors be reactivated after autoinhibition by phosphorylation?
phosphorylation of the inhibitory tail domain by the kinases MPS1 and CDK1-cyclin B, which results in processive motility on MTs
What is the lifecycle of kinesin motors?
- in the absence of cargo, kinesin motors are autoinhibited
- during transport, motor-cargo complexes must navigate MT tracks
- at the destination, cargo release is coupled to inactivation of the motor
What is dynein?
a minus-end motor (only 1 motor, kinesin has 5)
What is dynein made up of?
- 2 heavy chains
- 2 intermediate chains
- 2 light intermediate chains
- 2 light chains
What are the 3 domains of each dynein HC?
- head (motor) domain with ATPase activity
- linker domain where MTs bind, and movement is generated
- tail domain involved in dimerisation and cargo binding
What are the intermediate chains in dynein?
a scaffold that links the HC with the LC and binds dynactin
What do the light intermediate chains in dynein do?
bind adapter proteins (attached to HC)
What are the light chains in dynein involved in?
the assembly of the dynein complex and cargo interactions
What drives dynein motor action?
ATP hydrolysis
What are the 5 steps of the dynein ATP hydrolysis cycle?
- ATP binds to site at AAA1and releases the motor from the MT
- remodelling of the linker to induce a bend and subsequent ATP hydrolysis
- the motor diffuses to a new site and rebinds
- the linker straightens during a power stroke to produce force which moves the whole dynein molecule forward and can induce tension between the 2 motor domains
- ADP release returns the motor to the beginning of the cycle (state 1)
What do dynein adapters do?
regulate dynein motility
What is dynactin?
an essential dynein adapter that is a complex of 23 proteins
What are the functions of dynactin?
- attaches cargoes to dynein
- activates dynein motility
- targets dynein to plus end of MT tracks
- coordinates transport between dynein and kinesin motors
What are the 3 main components involved in directing motors and cargoes to their destinations?
- the AIS and motor targeting
- motors exhibiting intrinsic directionality
- cargo binding
What can switching cargoes do to motors?
cause them to switch direction
What is the new material to support during development?
- axon and dendrite outgrowth
- formation of synapses to form neuronal networks
What is polarised trafficking required for in mature neurons?
- addition of new proteins (synaptic plasticity)
- degradation of faulty proteins
- transport of signalling molecules from synapse to nucleus
Why does transport deliver biomaterials for neurite growth?
there is a rapid expansion of size
What is synaptogenesis?
formation of synapses
What is synaptogenesis essential for?
polarity and function (neurotransmission)
When do synapses form?
upon axon-dendrite contact
What are the 3 steps of synaptogenesis?
- migration
- contact
- synapse formation
What establishes and maintains synapses?
kinesin transport
When are cargoes deposited?
when the axons and dendrites make contact i.e. when synapses form
What are the 2 transport vesicles in axons?
- synaptic vesicle precursor transport vesicles e.g. KIF1
- piccolo-bassoon transport vesicles e.g. KIF5
What is the main transport vesicle in dendrites and what do they do?
PSD95 containing vesicles that form postsynaptic densities
What do the synaptic cell-adhesion molecules neurexins and neuroligins do?
- connect pre- and postsynaptic neurons at synapses
- mediate trans-synaptic signalling
- shape neural network properties by specifying synaptic functions
What does membrane fusion do?
deliver cargoes to their destination
What happens after vesicles are released from cargo?
they undergo tethering
What does dynein transport?
biomolecules back to the cell body for signalling and degradation
What can problems with neuronal transport cause?
synapse loss and axon degeneration
What can the APP family do?
- both negatively and positively modulate main transmission systems
- tune neuronal excitability by regulating intracellular calcium levels
What can happen after focal blockages of axonal transport?
swelling then degeneration of the distal axon
Give examples of axonal transport disruption and neurodegeneration
- HD
- AD
- PD
What does synaptic loss correlate to?
disease severity i.e. cognitive decline correlates to a decrease in the number of synapses in the brain
What happens in prodromal AD?
- circuit disruption
- mild synapse loss
- amyloid fibril forms on excitatory neuron
What happens in late-stage AD?
- hyper excitability
- inflammation
- synapse loss
- loss of inhibitory tone
- deficits in autophagy
What is a classic protein affected by reduced levels of motor proteins in AD patients?
synaptophysin
What are Aβ oligomers?
highly neurotoxic oligomers that can further aggregate to form soluble protofibrils (large oligomers) and then insoluble β-pleated sheets forming amyloid fibrils and plaque
What are the 7 stages leading to neurodegeneration?
- initiating factors e.g. ageing, lifestyle, toxins
- pathological proteins e.g. abnormal tau, Aβ oligomers
- aberrant kinase and phosphatase activity e.g. PP1, GSK
- impaired fast axonal transport
- synaptic dysfunction and axonal degeneration
- functional impairment e.g. memory loss, cognitive defects
- neurodegeneration