Lecture 18 Endocytosis And Nucleo/Cytodolic Transport Flashcards
Clathrin mediated endocytosis
Movement of membrane impermeable molecules into cell
Internalisation of surface proteins
Endocytosis
Internalisation of the plasma membrane
Used to internalise molecules e.g. nutrients
Used to control cell surface proteins such as receptors
Catherine mediated endocytosis is mediated by clathrin and adaptors
LDL particle internalisation (an example of endocytosis)
1) dietary lipids and cholesterol are transported in low density lipoprotein (LDL) particles
2) LDL binds to LDL receptor on phospholipid membrane
3) adaptor proteins recruit clathrin leads to formation of clathrin coat
4) coated pit/vesicle
5) vesicle un coats to form early endosome
6) early endosome fuses with late endosome
7) matures into lysosome containing fatty acids, aminos and cholesterol
8) receptors return to cell membrane
Familial Hypercholesterolemia (FH)
Caused by mutation in LDL receptor or ApoB
Clathrin cage assembly
Vesicle formation involves coating
Endocytosis > clathrin coat
Clathrin attaches to membrane via adaptor
Drives membrane deformation by assembly of curved cages
Once vesicle is formed adapter proteins and clathrin detach
Clathrin cage structure: triskelions
Clathrin cage structure formed from multiple hexameric complexes called triskelions
Made of 3 heavy chains stabilised by 3 light chains
Heavy chain tips are binding sites for assembly proteins
Endocytotic vesicle release by dynamin
Dynamin and associated proteins assemble around neck of bud forming a dynamin collar.
Then probably ‘squeezes’ neck of bud by confirmational change due to hydrolysis of GTP until two sides of the bud come together and fuse
Dynamin “shabire” mutant
Temperature sensitive mutation that causes paralysis in drosophila. Causes formation of buds that do not pinch off.
The mutant is non- functionally temp. Sensitive - blocks synaptic vesicles recycling resulting in neurotransmitter release blockage.
Reversible - when cooled down processes restart
V and T SNARE rol in targeting to correct compartment and membrane fusion
SNARE - sensitive factor attachment protein receptors
V-SNAREs - present on vesicle
T-SNAREs - present on target
Specific vesicle SNAREs bind to specific target SNARES, docking and fusion
SNAREs role in fusion
V and T SNAREs pull vesicle into contact with target membrane to allow fusion
V/T SNARE tight complex
SNAREs are unstructured until they bind to each other forming a 4 helix bundle
Brings the 2 membranes into close contact excluding H2O and allowing
fusion
SNARE mediated fusion is via first hemifusion intermediate - fusion of outer membranes and then internal membrane to complete fusion.
E.g. V-SNARE synaptobravin on synaptic vesicle binds to T-SNARE syntaxin on nerve plasma membrane
Nuclear envelope
Separates nuclear/cytoplasmic compartments
Nuclear pore mediates movement of
1) proteins to nucleus
2) mRNA to cytoplasm
Transcription in nucleus / translation on outer side of nuclear envelope and rough ER (separated unlike in bacteria)
Nuclear pore
Contains nuclear pore complex a lot of proteins working together to form a “leaky gate”
Nucleopore has 8 fold rotational symmetry
Nucleoplasmic ring has a sieve like structure covering the inner side preventing egress of unspliced mRNA
Nuclear transport is receptor mediated
Uses internal non-cleaved signal sequence NLS
Nuclear localisation signal attached to prospective nuclear protein
Attaches to nuclear pore receptor and passes into nucleus
Where receptor detaches and returns to cytosol
Cargoes contain amino acid import/export signals
Nuclear import:
For protein import
Uses Nuclear Localisation Signal (NLS)
NLSs are short and basic
E.g. NLS of large T-antigen of the virus sv40:PKKKRKV
Nuclear export
For protein export
Uses Nuclear Export Signal (NES)
E.g. the leucine rich signal of the HIV Rev Protein LQLPPLERLTL
May be
monopartite - have 1 basic amino acid cluster
Or
Bipartite - have 2
