Lecture 15: Immune System Protein Trafficking + viruses Flashcards
MHC stands for ___
Major Histocompatibility Complex
MHC are ____ on ___
Oligomeric protein complexes on the cell surface
MHC is ___ meaning that it has multiple alleles
polymorphic
MHC binds to ____ (___ amino acids)
wide variety of peptide antigens (8-24 a.a)
MHC (general) presents antigens to ____
T-cells
antigens to MHC I cause ___ while antigens to MHC II cause ___
cell death
T-cell stimulation
MHC complexes rich the cell surface with ___ using ____
bound peptide antigens
different trafficking pathways
class MHC I description
a-chain (transmembrane heavy chain)-polymorphic
->a1 +a2 =peptide binding groove
b-microglobulin - non polymorphic/ soluble
class MHC II description
a-chain + b-chain (transmembrane region)- 2 chains
a1 +b1 =peptide binding groove (polymorphic)
MHC I is present only in ___
nucleated cells (not in blood cells) in vertebrates
MHC II is present only in ____
antigens presenting cells (ex. dendritic cells, b-cells, macrophages)
MHC I presents antigen to ___
cytotoxic T-cells
MHC II presents antigen to ___
helper + regulatory T-cells
source of peptide antigen (MHC I)
-proteins made in cytoplasm
source of peptide antigen (MHC II)
endocytosis PM + extracellular proteins
If antigen on MHC I is foreign then ____
the cell is targeted for destruction
receptor on cytotoxic T-cell that recognizes MHC I
CD8
general structure of MHC I
heterodimer
a-chain of MHC is ____ membrane protein
Type-1
complete assembly of MHC I occurs in ___
ER
a-chain of MHC I associate with __, __, ___ before/during assembly in ____
calnexin*, calreticulin and ERp57
ER lumen
a-chain of MHC I associate with B-microglobulin
when?
-shortly after synthesis (2 min)
peptide binding groove of MHC I protein faces ___
lumen of ER
antigens (peptides) for MHC I are transported from ____ into ____ by ____
*cytosol into ER lumen by TAP
TAP is a ____ in ER membrane, ____-ase, & ____
Transporter
ATP-ase
heterodimer (TAP-1,TAP-2)
transport of antigens for MHC I into ER lumen is a ___-dependent process
ATP
peptides for MHC I (viral) are derived from ____
proteolysis of newly synthesized viral proteins in
cytosol by Proteasome
Tapasin (Tsn) is an _____associated with
ER membrane glycoprotein
TAP
Function of Tapasin
Facilitates peptide loading onto MHC I
___ motif on Tsn retains unloaded MHC I in ER
KK
Tsn has a ____(ER-retention) motif
dilysine
TAP has a ____(NBD) for __ binding
nucleotide binding domain
ATP
Tsn and fully loaded MHC I
Tsn releases MHC I to move to golgi and then cell surface
MHC I acts as a ___ for T-cell
a window
cytotoxic cell needs very little ___
cytoplasm (mostly nucleus)
In un-infected cells, most MHC I complexes are occupied by _____
peptides derived from cellular proteins (mostly defective proteins from ribosomes, some retirees)
In infected cells, most MHC I complexes are occupied by _____
viral peptides
state of MHC when cell is not fighting infection
-bound but with host proteins
receptor on helper T-cell that recognizes MHC II
CD4
MHC II a and B chain are both ___
type 1 membrane glycoproteins
peptide binding groove of MHC I binds peptides with ___ a.a
8-10
peptide binding groove of MHC II binds peptides with ___ a.a
12-20
assembly of MHC II begins in ____ and requires ___
ER
chaperones
invariant chain facilitates transport of _____
MHC II complexes to endosomes
invariant chain prevents ___
binding of peptides to MHC II in ER (don’t want MHC I peptides)
invariant chain is a ____
nonpolymorphic transmembrane protein(homotrimer)
invariant chain stimulates ___
exit of MHC II heterodimers from ER
invariant chain _____ peptide binding groove of MHC II by _____ in _____
is removed
proteolysis
endosomes
peptides of MHC II are derived from ___
degraded endocytosed material (bacteria, viruses) from cell surface
Proteasome _____ required to generate peptides for binding to MHC II
is not
After removal of invariant chain, MHC II binds to peptide in _____ which ____ complex
endosomes
stabilizes
invariant chain contains ___ motif in ____
dileucine (LL/ML)
cytoplasmic domain
Viral immune evasion strategies
-Avoiding antibodies-changing of surface proteins (flu,cold,coronaviruses)
-Viral proteins that modulate host immune response
How do viral proteins modulate host immune response?
-Blocking expression of cytokines and prevention of
programmed cell death (apoptosis)
– Interfering with cellular immune response by disabling peptide
presentation or impairing NK cell function
Viral Interference with MHC I Function (Post-Translational): generation of peptides
EBNA-1 protein contains Gly-Ala repeats which interferes with proteasome (no peptides degrades so no assembly on MHC I and export from ER)
Viral Interference with MHC I Function (Post-Translational): peptide transport in ER
-competes for peptide-binding of TAP
-stabilizes TAP in conformation that it can’t bind to ATP
result: no peptides in ER, no assembly of MHC I, no export from ER
E3-19K is a ____
“immunosubversive” protein encoded by adenoviruses
adenoviruses are ___
small DNA viruses (often cause cold-like symptoms)
E3-19K has which motif?
KK motif in cytoplasm
E3-19K blocks the expression of ___ at _____ by preventing the ______
MHC I
cell surface
conversion of N-linked sugar into Endo H resistant forms
E3-19K binds to and retains MHC I in ___
ER
E3-19K also binds to ___ and prevents binding to ____
TAP
tapasin (peptide-loading)
CMV US3 and US10 have same function as ___ but don’t prevent ___
E3-19K
peptide loading
CMV US2 and US11 bind to ___ in___ and cause
MHC I
ER
dislocation
US11 localizes where?
in ER
proteasome inhibitor
ZL3VS
____inhibits US11-induced degradation of MHC (HC-heavy chain)
Blocking proteasome
US-11 leads to expression of ____
soluble, deglycosylated MHC I HC
Differential centrifugation refresher
-Start from cell homogenate
-Spin at different speeds (g force) for various times
-Obtain several fractions that are enriched in cell components (e.g. membranes) of various sizes
Differential centrifugation 1000 x g pellet
-whole cells, nuclei, cytoskeletons
Differential centrifugation 10,000 x g pellet
-mitochondria, lysosomes, peroxisomes
Differential centrifugation 100,000 x g pellet
microsomes, small vesicles
HIV Nef protein has multiple functions
- Not required for replication
- Involved in immune deficiency
Nef protein operates ____
post ER/Golgi
Nef-induced degradation of ____ is blocked by inhibitors of ____
mature MHC-I
acidic degradation
Lactacystine (Lact) inhibits____
proteasome
Bafilomycin (Baf A1) inhibits ___
proton pump in lysosomes
NH4Cl raises _____
pH of lysosomes
both bafilomycin +NH4Cl ___
disrupts function of lysosomes
HIV Nef causes _____
lysosomal-dependent degradation of MHC I (needs AP-1)
depletion of AP-1 effect on Nef-dependent degradation of MHC I
-inhibits degradation of MHC I(AP-1 moves MHC I from golgi to lysosomes)
______ interacts with AP-2 to accelerate endocytosis of CD4 and MHC-1, leading to ____
HIV-Nef
selective removal of MHC I from surface of presenting cell
interference of MHC II mostly occurs in ___
endocytic pathway
Papilloma viruses + MHC II interference
produces E5 protein: replace subunit of proton pump
-pH in endosomes/lysosomes increases
-Proteolysis of endocytosed material is blocked
-no peptide-loading of MHC II
-decreased cell surface
expression of MHC II