Lecture 9: Protein and Vesicular Transport

Question 1

where do proteins synthesised by ribosomes on the RER go

Answer 1

• nuclear membrane
• secretory vesicles
• peroxisome membrane
• packaged by Golgi to go to plasma membrane

Question 2

what tells the protein where to go

Answer 2

signal sequence

Question 3

what do proteins need to do to pass through the membrane

Answer 3

unfold and then refold when they’re through the membrane

Question 4

what are the proteins that make up the nuclear pore

Answer 4

nucleoporins

Question 5

what are the steps of nuclear import

Answer 5

1) protein’s nuclear localisation signal is recognised by importin, which binds to proteins within the cytosolic fibrils
2) importins carry the protein into the nucleus by disrupting the gel-like mesh of nucleoporins, making a channel
3) once inside, an interaction with Ran protein causes release of cargo proteins and returns importin back outside the pore

Question 6

what is Ran and how does it work

Answer 6

• GTP binding protein
• when bound to GDP, Ran binds to importin and takes it into the nucleus while bound to NTF2
• inside the nucleus, Ran GEF exchanges GDP for GTP
• this releases importin back to cytosol

Question 7

how does a mitochondrial protein get through the outer membrane

Answer 7

signal sequence recognised by import receptor protein, which will interact with a protein translocator in the outer membrane

Question 8

how does a mitochondrial protein get through both membranes to the matrix

Answer 8

• gets through the outer membrane
• travels along membrane to a spot where both membranes are in close proximity
• interacts with protein translocator on inner membrane is unfolded into primary structure and fed through membrane
• once inside the matrix, signal sequence is cleaved off and chaperones fold protein back into its final shape

Question 9

how are proteins transported to the endoplasmic reticulum

Answer 9

• protein translocator binds signal sequence during translation and threads polypeptide through as a loop
• signal sequence is cleaved off and left in membrane to degrade
• as polypeptide feeds through, chaperone BiP binds, ready to help fold protein back within the ER

Question 10

how do the ribosomes know they have to transport the protein to the ER

Answer 10

• as the ribosomes start manufacturing the protein, the signal sequence is recognised by signal recognition particles (SRPs)
• SRP binds to SRP receptor on surface of ER lumen
• signal sequence transferred across to protein translocator, starting protein import

Question 11

how are single-pass transmembrane proteins imported into the ER membrane

Answer 11

• signal sequence begins transfer into membrane
• when stop-transfer sequence reaches protein translocator, protein is stopped from being fed through
• signal sequence removed, releasing inner end of the sequence into the ER
• outer part remains outside ER and stop-transfer sequence becomes transmembrane domain

Question 12

how are double-pass transmembrane proteins imported into the ER membrane

Answer 12

• instead of the signal sequence being at the end of the amino acid sequence, it’s embedded within
• translocator protein keeps feeding it through until it hits stop-transfer sequence, and ejects protein into the membrane

Question 13

what is glycosylation

Answer 13

adding a branched oligosaccharide side chain to an appropriate asparagine

Question 14

what is glycosylation important for

Answer 14

• inflammatory response
• could enable viral immune escape
• promoting cancer cell metastasis
• regulating apoptosis

Question 15

what happens if a protein misfolds

Answer 15

• retained in ER by chaperones until it can be folded correctly, or exported into cytosol for degradation
• excess of unfolded proteins triggers the unfolded protein response
• increases size of ER, number of chaperones and decreases transcription

Question 16

how do vesicles travel from the ER to other places

Answer 16

• bud off ER and each level of Golgi membrane to gt to the Golgi apparatus
• bud off Golgi into early and late endosomes or lysosomes to get to plasma membrane
• exocytosed vesicles replenish lipids and cell surface receptors at surface

Question 17

what are the pathways of vesicular transport

Answer 17

• constitutive: unregulated, continued secretion of proteins from the cell
• regulated: secretory vesicles store high concentrations of protein and wait until an extracellular signal stimulates their secretion

Question 18

what are coat proteins

Answer 18

• proteins that aid in vesicle budding

- after budding, the coat is shed to expose lipids on the vesicle which permits fusion with the target membrane

Question 19

which coat proteins help transport from Golgi to lysosomes

Answer 19

clathrin and adaptin

Question 20

which coat proteins help transport from plasma membrane to endosome

Answer 20

clathrin and adaptin

Question 21

which coat proteins help transport from ER to Golgi

Answer 21

COPI

Question 22

which coat proteins help transport from Golgi to ER

Answer 22

COPII

Question 23

what is clathrin

Answer 23

• coat protein made up of 3 chains that form basket like cages
• involved in shaping the curvature of the budding membrane
• clathrin coated pits form on the membrane

Question 24

what is dynamin

Answer 24

• a GTP binding coat protein that assembles around the neck of budding vesicles to pinch it off
• squeezes lipids together until they fuse and flow into each other

Question 25

how is the protein recognised at the target membrane

Answer 25

• tethering proteins recognise Rab protein on vesicle and pull it towards the membrane
• v-SNARE binds to complementary t-SNARE on target membrane, which wind around each other to pull vesicle close to membrane
• squeezes away water molecules, allowing lipids to flow and form a continuous layer
• SNAKES unwind to be used again

Question 26

how do phagocytes engulf pathogens

Answer 26

• forms pseudopods which is the membrane expanding
• viruses can also bind to receptors on the cell membrane and be taken up by receptor mediated endocytosis
• end up in clathrin coated vesicles
• low pH allows release of viral genome into cytoplasm

Question 27

how are receptors degraded

Answer 27

• delivered to early endosome, then late endosome, then lysosome where they get destroyed and degraded
• important when the signal needs to be turned off

Question 28

how are receptors recycled or transcytosed

Answer 28

• ligand dissociates in early endosome
• can return to same membrane they came from (recycling)
• or sometimes go to a different domain (transcytosis)

Question 29

what is autophagy

Answer 29

breakdown of the cell’s own proteins and organelles by enclosing them in a membrane that fuses with the lysosome