Week 5 - yeast in understanding cell membrane trafficking pathways

Question 1

What is glycosylation?

Answer 1

modification of an organic molecule, especially a protein, by addition of a sugar molecule.

proteins are modified by oligosaccharide addition in the ER and processing in the Golgi.

Question 2

What are the advantages and disadvantages for yeast as a model organism?

Answer 2

advantages:
- amenable for genetics studies
- entire genome sequence is known
- cheap and easy to grow
- limited gene diversity
- fundamental pathways are conserved.

Disadvantages:
- informative about multicellularity as limited cell to cell contact.
- small so high resolution images of compartments is difficult.

Question 3

In the Novick and Schekman experiment - what did they believe would happen if proteins could not be secreted?

Answer 3

the cell would increase in density as the vesicles carrying the proteins would accumulate, instead of being secreted.

Question 4

What are Sec genes?

Answer 4

Sec = stands for secretion

• involved in protein synthesis and secretion in eukaryotic cells.
• mainly for synthesis and transport of proteins to the endoplasmic recticulum.

Question 5

What is endocytosis?

Answer 5

Endocytosis is when the plasma membrane invaginates into the cell, producing a vesicle which can then fuse with endosomes and enter the end-lysosomal membrane system

Question 6

What are the stages in the endocytic pathway?

Answer 6

1) Plasma membrane to endocytic vesicle

2) endocytic vesicle to early endoscope

3) early endosome to late endosome (MVB) or recycling to the plasma membrane.

4) late endosome to Golgi/vacuole.

Question 7

What is the major function of the lysosome (vacuole)?

Answer 7

for degradation of extracellular material taken up by endocytosis

(and some intracellular components by auto-phagy).

Question 8

What does MVB stand for and what is it?

Answer 8

MVB = multivesicular body

• are found in the late endosome
• formed from invagination of the MVB membrane, which makes vesicles that are separate from the main lumen.

Question 9

What are the 4 possible trafficking pathways from the late Golgi?

Answer 9

1. to the plasma membrane
2. to the early endosome
3. to the late endosome/MVB
4. to vacuole.

Question 10

The protein CPY (carboxypeptidase) has been used in yeast genetic screen to be able to identify distinct trafficking compartments.

What properties of CPY are central to its use in the screens?

Answer 10

It is glycosylated and proteolytically processed in different compartments

Question 11

What are the steps in the CPY pathway?

Answer 11

In summary, CPY moves from the ER to the Golgi, is sorted by Vps10, transported by cytoplasmic factors, and then cleaved into its mature form in the vacuole. Vps10 is then recycled back to the Golgi.

1. CPY is made prepro and moves through the ER to the Golgi.
2. In the late Golgi, CPY is recognized by a receptor called Vps10, which sorts it to its correct location.
3. The transport of CPY requires cytoplasmic factors: clathrin and two adaptors called Gga1 and Gga2.
4. CPY separates from Vps10 in the late endosome/MVB and moves to the vacuole, where it is cleaved into its mature form.
5. Vps10 is returned to the late Golgi through a specific signal sequence in its protein sequence (YSSL, FYVF).