Lecture 6 - Research Analysis I - Cross presentation Flashcards
Compare the origin of Ag present in the context of MHC I and MHC II
MHC I:
Ag from cytosol
MHC II:
Ag from endosomal compartments
What is the inherent problem with priming CTL responses against viruses that do not infect DCs?
To prime CTL responses, the Ag needs to be presented in the context of MHC I
For this to occur, the Ag must be present in the cytosol of the APC
If the APC is not infected, how can this Ag come to be in the cytosol
→ cross-presentation
Outline the various hypotheses of mechanisms of cross presentation:
• Endosomal pathway
• Cytosolic pathway
What are the problems with these hypotheses?
- Endosomal pathway
• Ags are processed in endosomes
• Peptides are loaded on MHC I molecules in endosomes
• MHC I molecules have been recycled from the plasma membrane
Problem:
• Unclear whether endosomal proteases can generate the same antigenic peptides that are generated in the cytosol
• Unclear whether ‘good quality’ MHC I:peptide complexes can be formed in the absence of the PLC
- Cytosolic pathway
• Endocytosed Ags are transferred to the cytosol
• Ags now can access the canonical MHC I presentation machinery
Problem:
• How do the Ags move from the endosome into the cytosol? (some transporter we don’t know about)
• Do we know if the proteasome is involved? (yes)
Describe Guermonprez’ paper on cross presentation
- Proteasome involvement
- Phagosome recruitment of ER components
- Proteasome involvement
Used a proteasome inhibitor (lactacytin)
Found that this inhibitor prevented the cross presentation of Ovalbumin
Implication:
• The proteasome is involved in cross-presentation
• (negates endosome pathway of cross-presentation)
- Phagasome composition
Used kinetic analysis
Demonstrated that the phagosome recruits ER components:
• Sec61
• TAP
• PLC components: Tapasin, ERp57, Calreticulin
Describe the implications of Ackerman and Cresswell’s paper on cross-presentation
- TAP involvement
- Sec61 involvement
- TAP involvement
ICP47 is a TAP inhibitor
ICP47 was endocytosed and inhibited TAP, blocking MHC I Ag presentation
Implication:
• ICP47 moved from endosome to cytosol
- Sec61 involvement
Inhibition of Sec61 was shown to block cross-presentation of OVA
Inhibition of Sec61 did not block presentation of SINFEKL
SINFEKL is a small epitope of OVA that can bind directly to MHC I on the surface of cells
Implication:
• Sec61 involved in cross-presentation
Describe the implications of Houde’s paper on cross-presentation
- Transfer of protein from endosomes to cytosol
- Proteasome involvement
- Transfer of protein from endosomes to cytosol
Used fluorescently tagged OVA
OVA was endocytosed by cells
60 mins after endocytosis, OVA was observed in the cytosol
Implication:
• Endocytosed OVA was able to move from endosome to cytosol
- Proteasome involvement
Demonstrated that ubiquitinated proteins converge around phagosomes
These ‘ubiquitinated proteins’ are substrates for the proteasome
Implication:
• Proteasome located near the phagosomes
What are the classical functions of Sec61?
Used in all cells as ‘Translocon’:
- Enables co-translational transfer of nascent polypeptides into lumen of ER
- Enables exit of misfiled polypeptides from ER lumen into cytosol for ubiquitination and degradation
Describe the ‘Hybrid endosomal-ER compartment’ hypothesis for cross-presentation
What are the components of this hybrid compartment?
What would this allow?
“Ergosome”
These compartments have components of both endosomes and ER compartments:
• Sec61
• TAP
• PLC
This hybrid is formed through the fusion of blebs from ER and the endosomes
These components respectively enable:
a. Sec61
• Transfer of endocytosed Ags to cytosol for proteasomal processing
b. TAP
• Import of antigenic peptides into lumen of hybrid compartment
c. PLC
• Assembly of ‘good quality’ MHC I:peptide complexes
