Lecture 5 - Antigen Presentation II Flashcards
In general, which cells can capture Ag from the environment and present it?
Professional antigen presenting cells
Describe the overall process of Ag presentation in the context of MHC class II
- APC endocytoses Ag
- Endosomal proteases (cathepsins) degrade the Ag
- Ag peptides are contained in endosomal compartments
- MHC II molecules are synthesised in the ER
- Trafficking of vesicles w/ MHC II from ER to endocytic compartment
- MHC II molecules bind the Ag peptides
- MHC II-peptide complexes are transported to the plasma membrane
What is the ‘goal’ of MHC II presentation?
To display in real-time on the plasma membrane a sample of all the proteins contained in endosomal compartments of the cell
When does Ag presentation on MHC II occur?
Constitutively:
In the absence of infection, only peptides derived from ‘normal self’ proteins are presented
Upon infection or cellular transformation ‘altered self’ proteins are also presented in conjunction with ‘normal self’ protein derived peptides
What degrades proteins in endocytic compartments?
Endosomal proteases called cathepsins
Describe the mechanisms of endocytosis that occur
What does each result in the formation of?
What is each process dependent on?
Describe the specificity of each mechanism
Which cells are capable of each mechanism of endocytosis?
Which mechanisms are most important in MHC II presentation?
- Macropinocytosis
• ‘Drinking’ a large volume
• Actin dependent (to bend the plasma membrane)
• Non-specific
• Results in the formation of a macropinosome
• DCs - Phagocytosis
• ‘Eating’ a large particle (bacterium, protozoans, cells)
• Actin dependent
• Triggered by a surface receptor (specific)
• Results in formation of a phagosome
• DCs, Macrophages - RME
• Capture of a soluble molecule
• Clathrin-dependent
• Specific: triggered by a surface receptor
• Results in the formation of a micropinosome
• DCs, Macrophages, B cells
also:
4. Pinocytosis
• 'Drinking' a small volume
• Clathrin-dependent
• Non-specific ("fluid phase")
• Results in the formation of a Micropinosome
- Autophagocytosis (?)
• Cytosolic proteins are transferred to endosomes
• Mechanism unclear
• Results in the formation of an autophagosome
Most important:
• Macropinocytosis
• Phagocytosis
• RME
Which mechanisms of endocytosis can the following cells perform:
• DCs
• Macrophages
• B cells?
DCs:
• Macropinocytosis
• RME
• Phagocytosis
Macrophages:
• RME
• Phagocytosis
B cells:
• RME
Which receptors do each of the following cells use to perform RME:
• DCs
• Macrophages
• B cells?
DCs:
• Mannose receptors
• FcRs
etc.
Macrophages:
• Mannose receptors
• FcRs
etc.
B cells:
• Only surface Ig
Describe the different aspects of the endocytic pathway
Describe the biochemical changes that occur along the endocytic pathway
- Early endosome
•The immediate product of endocytosis
•Contents largely materials from extracellular environment - Late endosome
•Contents are more ‘cellular’, and less ‘extracellular’ - Lysosome
•Final destination of materials in endosomes
•Degradation of materials into smaller components that may be recycled and used again by the cell
As the endosome ‘matures’, contents become increasingly:
• Acidic (pH decreases)
• Reducing (sulphide bonds destroyed)
• Richer in proteases
Describe the process of degradation of endocytosed protein
– Denaturation –
- Acidification:
• H+-pump ATPases lower the pH of endosomal compartments - Reduction of disulphide bonds
• GILT cleaves S-S bonds
• GILT: gamma interferon lysosomal thiol reductase
Cleavage of disulphide bonds plus low pH results in the denaturation of proteins → more accessible to proteolysis
– Proteolysis –
By endosomal proteases, known as Cathepsins (Cts + letter; e.g. Cts D)
Describe the features of cathepsins
• Abundant in all cells (not just APCs) → indicates general role in turn-over of endogenous membrane proteins and endocytosed extracellular proteins
Different active sites:
• Cysteine (Cts B, L, S)
• Aspartic acid (Cts D, E)
• Serine (Cts G)
Variable optimum pH
Rather nonspecific and redundant
• Different cathepsins will do the same thing (i.e. cleave at the same location)
Can be secreted for extracellular proteolysis
Describe the route that MHC II molecules take before they are loaded with peptide
- Synthesised in the ER
- Ii (invariant chain) binds to peptide binding cleft of MHC II in ER (to prevent binding of other ER components)
- Specific Ii sequence on cytoplasmic tail tags MHC II for transport to the endocytic route (not to the plasma membrane like MHC I)
- Zipper region of Ii is cleaved by an unknown mechanism, releasing three αβ dimers, each bound to Iip10
- Cathepsin S cleaves Iip10 at remove transmembrane tail (only CLIP still bound to αβ dimer)
- Chaperone HLA-DM interacts with αβ-CLIP, resulting in an ‘open conformation’ of the αβ dimer
- CLIP substituted by an antigenic peptide
8a. Preformed peptide binds to MHC II binding cleft
8b. Epitope capture:
• Longer polypeptides bind to the MHC II binding cleft, then are further trimmed down to size
- Complexes trafficked to the cell surface
- Remain on cell surface until the peptide dissociates from the MHC II binding cleft
Describe the peptide binding cleft of MHC II molecules
Open on both sides (“hot dog”)
Can bind peptides of any size
(Unlike MHC I, which binds peptides 9 aa in length)
Describe the structure of Ii
Transmembrane: embedded in the ER membrane, tail on the inside of the ER
Two regions:
- “Zipper” region
• Promotes formation of a trimer (three Ii) - CLIP region
• Occupies the peptide binding cleft of the αβ dimer
• CLIP is promiscuous: fits the peptide binding cleft of all MHC II allotropes
What are CIIV and MIIC?
CIIV:
• Early endosome that contains MHC II molecules
MIIC:
• Late endosome that contains MHC II molecules
