Lecture 12: Vesicular Transport within Golgi and Endosome

Structure of Golgi Apparatus

cis Side - ER facing side, trans Side - plasma membrane facing side

cis Golgi network (CGN)

cis cisterna, medial cisterna, trans cisterna

trans Golgi network (TGN)

what does exit signal do?

Cargo that needs to be transported has exit signals. Exit signal proteins are often packaged into vesicle to be transported

Which protein bound by cargo receptor?

soluble proteins bound by cargo receptors, which binds to adaptor protein of the inner COPII coat

What binds the transmembrane protein?

bound by COPII coat

How does cargo without an exit signal get packaged into vesicle?

diffusion - if concentration is high, can be pushed in

Vesicles that bud from exit site…

has COP II coat, form a vesicular tubular cluster by fusing with each other (this moves to golgi apparatus). NSF, using ATP, untangles v-SNARE and t-SNARE on its vesicles → tangle v-SNARE and t-SNARE on opposite vesicles → fuse together

What coated vesicles does Retrieval (Retrograde) transport use?

COPI coated vesicles (vesicles from vesicular tubular clusters and Golgi go to ER)

What do the retrieval proteins have?

has ER retrieval signals

Soluble ER proteins have KDEL (retrieval signal) → which needs to be bound by KDEL receptor for the protein to be brought back

What signal does KDEL receptor have?

some ER membrane proteins have a retrieval signal.

for KDEL, it has KKXX at C terminus → binds to the COPI coats, which is packaged into vesicles

KDEL Receptor and Mechanism of Retrieval of ER-Resident Proteins

KDEL cycles between ER and Golgi

ER: higher pH, lower affinity for KDEL (releases KDEL)

Golgi: lower pH, higher affinity for KDEL (grabs onto KDEL to bring back to ER)

Different mechanism for Retrograde Transport

1. Different transport rates: some cycle but transport to ER at a slower rate (so will accumulate in Golgi)

2. Chunks of proteins retained in resident compartment: no space to come in

Transport of protein from CGN to TGN basic info

vesicles are kept close to Golgi cisternae by tethering proteins

Processing of N-linked oligosaccharides occurs int eh Golgi

Transport of protein from CGN to TGN - 2 models

both uses COPI coated vesicles

• Vesicular Transport Model: COPI coated vesicles move forward on one Golgi cisterna to the next (faster)

• Cisternal Maturation Model: “everyone gets promoted” returning uses COPI vesicles (slower)

What is the cargo when transporting TGN to late endosome?

lysosomal hydrolases → needed for lysosome function and for degradation of macromolecules, synthesized in the ER, processed in Golgi, active at acidic pH

TGN vesicles going to late endosome what happens?

The cargo is coated in clathrin → transported to late endosome → mature into lysosomes

What happens when extracellular material gets into the cell via vesicles? (lysosome)

Joins early endosome (which has digestive enzymes) → gobble more things up = late endosome → eventually gets degraded, and we call this organelle lysosome

What happens to late endosome, after it becomes lysosome?

It will still have hydrolase left → fuse it to endosome and go through process again

Detailed Mechanism for Lysosome being made

• N-linked oligosaccharide added to ER

• mannose is phosphorylated by…

  • Phoso-N acetyl glucosamine is added

  • N-acetyl glucosamine is removed → phospho left

  • = M6P

• M6P receptor in TGN package lysosomal hydrolase

• Lysosomal hydrolases relased into late endosome

Affinity of M6P receptor

Lysosome: at low pH = decreased M6P receptor affinity = release cargo

Golgi: at high pH = increased M6P receptor affinity = grab on to send it to lysosome