Lecture 9. Synthesis and Packaging of Signaling Molecules for Release Flashcards
How does the secretory system function ?
Through the use of vesicular transport
What is vesicular transport ?
When transport vesicles carry soluble proteins and membranes between compartments
What did Pallad show us ?
How proteins move through at a subcellular level
What is sec 1 protein essential for ?
Vesicles fusing with a membrane which is called Munc 18
What can vessels with sec 1 do ?
Carry a full cargo intended for the cell surface but cannot merge with it and releases their contents
What does sec 7 do ?
Causes a huge swelling in the golgi network that prevents vesicles from forming
What happened when sec 1 and sec 7 are crossed ?
The swellings still formed but vesicles were not produced.
This tells us that vesicles production was downstream or later in the process than swelling formation
What do proteins destined for the membrane or secretory pathway have ?
A signal sequence on the nascent polypeptide
What does having a signal sequence on the nascent peptide target the proteins for ?
Binding with the protein signal recognition particle
How many particles does the signal recognition particle contain ?
6
What is the function of the signal recognition particle ?
Binds to the signal sequence and take it to the endoplasmic reticulum
What happens once the signal recognition particle has bound the signal sequence ?
It is recognised by the signal recognition particle receptor protein which is in the membrane of the rough endoplasmic reticulum
What happens when the signal recognition receptor protein complex binds ?
It directs the nascent protein into the lumen of the endoplasmic reticulum
Where does the signal peptide go through ?
The translocator complex
What happens when the signal peptide goes through the translocator complex ?
The signal peptidase cleaves off the signal peptide which is left in the membrane and the mature protein is cleaved off and liberated into the endoplasmic reticulum
What is the function of sugars that bind to the protein as it emerges into the endoplasmic reticulum ?
Regulate protein folding
What are the chaperone proteins that recognise the sugar binding ?
Calnexin and calreticulin
What are calnexin and calreticulin specialised to do ?
Recognise when the sugars are being cleaved off and therefore retain the protein in the endoplasmic reticulum until the protein is correctly folded
What is the signal that protein folding is complete ?
Once the sugar residues have been cleaved off
What do exit signal proteins do ?
Recognise the released proteins with some specificity and begin to form vesicles
What is the function of coat protein complex coated vesicles ?
They are structurally important in organising the form of the vesicle
What can the vesicle coated with COPII do once it forms the vesicular tubule structure ?
Can bud off
How does the vesicle coated with COPII travel ?
In an antergrade fashion along the microtubule
What do vesicles coated with COPII use to walk and carry the cargo ?
Motor proteins like kinesin
What is vesicle coated with COPII targeted according to ?
What they are coated in to move towards the golgi network
In the golgi apparatus, what are vesicles sorted into ?
Cis-cisterna, trans-cisterna and the trans-golgi network
What labels the layers in the stack in the golgi ?
Different enzymes or substances
What happens to the vesicles after they have gone through several processes in the golgi?
Proteins are sorted into new vesicles which bud off from the trans golgi network to the final destination in the cell
How does insulin messenger RNA go from the ribosome to the endoplasmic reticulum ?
Translator complex
What is insulin synthesised as ?
Pre pro insulin
What does pre pro insulin have that targets it to the signal recognition complex ?
A signal sequence
What part of the pre pro insulin gets cleaved off in the endoplasmic reticulum ?
The signal sequence
Once the signal sequence is cleaved from the pre pro insulin how many amino acids remain ?
76
What happens after pre pro insulin cleavage ?
It starts to fold but three disulphide bonds form which keep the a and b parts of the peptide together
What is between the a and b peptide ?
The c peptide
What is the structure of the c peptide ?
It remains continuous with the a and b peptides because of the pair of basic amino acids at both ends
What does endopeptidase do ?
Cleaves at two positions and release the c peptide
What does carboxypeptidase do ?
Cleaves off the basic amino acids. This occurs in the secretory vesicle
What is the result of carboxypeptidase cleavage ?
A and b chain dimer with disulphide bonds
Where is insulin initially packaged ?
Into immature secretory granules emanating from the trans golgi network
What is step 1 on granule maturation ?
Acidification of the granule lumen via the activation of proton pumps lower the luminal pH.
This is the optimal pH for activation of proteases and consequent conversion of proinsulin to insulin and C-peptide through proteolysis by endoproteases, followed by the trimming of the carboxyl termini carboxypeptidase E
What is step 2 on granule maturation ?
In the presence of calcium and zinc, it forms hexamers and effectively crystallises forming electron dense cores
What is step 3 on granule maturation ?
Fusion using SNARE complexes, vesicles fuse with the target membrane. Through synaptotagmin binding of incoming calcium, vesicle contents are released
What does mature insulin wait for to be released ?
Metaboli signals to exocytosed from the cell into circulation
What are the two stages of beta cell release of insulin ?
- Rapidly released in response to increased blood glucose levels
- There is a sustained slow release of newly formed vesicles triggered independently of sugar
How is insulin released from secretory cells ?
Calcium dependent exocytosis
How is calcium influxed ?
Comes through voltage dependent calcium channels
How is calcium released for the endoplasmic reticulum ?
By ryanodine receptors
How is insulin secreted ?
Via the fusion of a vesicle filled with a plasma membrane
Why is the fusion of a vesicle with a plasma membrane electron dense ?
They are basically crystallised hexamers of insulin, which are cross-linked by aldehyde fixation
What is a neuron ?
A highly specialised structure with branched dendrites for receiving signals from other neurons and a long highly specialised axon which conveys messages from this cell to another
What is the axon end called ?
A presynaptic terminal
What does the presynaptic vesicle contain ?
Secretory vesicles
What is the long stretch of axon between cell bodies and the presynaptic terminal coated with ?
Myelin
What is the function of myelin ?
Insulates the axon and allows electrical conductance to flow more rapidly down the axon
What makes myelin ?
Oligodendrocytes
What are astrocytes responsible for ?
Neurotransmitter reuptake, insulation/isolation, migration, synaptic plasticity
Where are astrocytes found ?
Brain
What are microglia ?
Immune function scavengers with possible roles in synaptic regulation
What is the neuron composed of ?
Cell body, axon and synapse
What is neurotransmission?
The conversion of an electrical signal to a chemical signal down the axon, that allows the signal to pass from one cell to the next
Where does the conversion of the electrical signal to a chemical signal occur ?
The synaptic cleft
Where is acetylcholine released from ?
The vagus nerve
Where does acetylcholine act on ?
Receptors in the heart
How does the noradrenaline speed up the heart rate ?
Through the accelerator nerve
Where is the post synaptic density found ?
Specialised structure on a spine on a cell
What does the post synaptic density receive ?
The synaptic enervation
When do post synaptic density release the neurotransmitter ?
With elevated levels of calcium
What is one of the key differences of the post synaptic neurons ?
They release substances that are not protenatious
What are some examples of neurotransmitters ?
- Biogenic amines
- Amino acid neurotransmitters
- Others
What is the four criteria for designation as a neurotransmitter ?
- Synthesis in neuron
- Verifiable release from neuron
- Effect on post-synaptic neuron via a specific post synaptic receptor
- Appropriate termination mechanism
What are catecholamines ?
Neurotransmitters that are derived from tyrosine
What is the structure of a catecholamine ?
The catechol ring structure and the amine group
How is DOPA formed ?
In the cytoplasm, tyrosine hydroxylase hydroxylates tyrosine by adding an OH to the catechol ring
How is dopamine formed from DOPA ?
DOPA decarboxylase decarboxylates DOPA
How is noradrenaline formed from dopamine ?
Dopamine beta hydroxylase adds a hydroxyl group to the beta carbon of dopamine
What is a major driving of parkinsons ?
Loss of cells that makes dopamine
Where are amine neurotransmitter synthesised enzymatically ?
In the cytosol
Where is amine neurotransmitters packaged into vesicles ?
Synaptic vesicles
What are the two key proteins that small vesicles are specialised by ?
- Vesicular neurotransmitter transporter
2. A vesicular ATPase which pumps H+ into the lumen
What are the two linked but distinct processes that are involved in the packaging of synaptic vesicles ?
- Vesicular ATPase pumps H+ into the vesicle against the concentration gradient and therefore require ATP hydrolysis
- A vesicular neurotransmitter transporter, transports neurotransmitters into vesicles against the concentration gradient by harnessing the power provided by the flow of H+ ions, down the concentration gradient out of the vesicle
What do all four types of neurotransmitter transporters contain ?
12 membrane spanning regions called transmembrane domains
What are the four types of neurotransmitter transporters ?
- VMAT
- VaChTs
- VGAT
- VGLUT
What are VMATs ?
Amine transporters that transport biogenic amines all positively charged at neutral pH and require outward movement of 2H+
How is vesicle packaged ?
Using neurotransmitter transporters
