Introduction
Class I histocompatibility molecules
play an important role in the immune response to intracellular pathogens
such as viruses. They function to "present" foreign antigens to cytotoxic
T cells in a process termed antigen presentation. Class I molecules are
cell surface membrane glycoproteins that consist of three subunits: a transmembrane
heavy chain (H chain), a soluble subunit termed beta2-microglobulin (beta2m),
and an 8-10 amino acid peptide (Figure 1).
Figure 1. Crystal structure of the soluble, extracellular portion of a class I histocompatibility molecule with the H chain depicted in green, beta2-microglobulin in yellow, and bound peptide in red. The H chain consists of three domains termed alpha1, alpha2, and alpha3. The peptide binds within a groove formed by the alpha1 and alpha 2 domains of the H chain.
In an infected cell, foreign proteins are degraded in the cytosol by the proteasome complex and peptide fragments are delivered into the endoplasmic reticulum (ER) by a transporter protein known as TAP. Within the ER, newly synthesized class I molecules bind to some of these peptides and the resulting class I-peptide complexes are exported to the cell surface. Here they are scrutinized by T cell receptors on cytotoxic T cells and, if the peptide presented by class I is recognized as "foreign", the infected cell is killed (Figure 2).
Figure 2. Within
the ER, newly synthesized H chains (green) associate with a membrane-bound
molecular chaperone termed calnexin (orange) and another protein ERp57
(purple). The calnexin-H chain complex then binds to the beta2m subunit
(green). At this point, calnexin may remain or it may be replaced by its
soluble homolog calreticulin. Yet another protein, Bap31 (blue), binds
at this stage. Subsequent association with the TAP transporter (orange)
occurs in an interaction that is bridged by another protein termed tapasin
(pink). This multi-subunit complex consisting of calnexin (or calreticulin),
H chain, beta2m, tapasin, TAP, ERp57 and Bap31 is known as the "peptide-loading
complex" or PLC. Following peptide binding to class I, the peptide
loading complex dissociates and the fully assembled class I molecule is
exported to the cell surface. Once at the cell surface, peptide-class
I complexes are scrutinized by T cell receptors on circulating cytotoxic
T cells. If the T cell receptor binds with sufficiently high affinity,
the infected cell will be killed.
Functions of components of
the antigen presentation pathway
A major focus of my laboratory
is to understand how the intracellular complex of peptide and class I molecule
is assembled. We discovered a
novel molecular chaperone known as calnexin that participates in the
folding and assembly of many nascent proteins including class I. We have
shown that calnexin, as well as a related protein termed calreticulin, function
to
prevent exit of incompletely assembled class I molecules from the ER
and also protect assembly intermediates from rapid intracellular degradation.
Furthermore,
calnexin and
calreticulin promote the assembly of class I-peptide complexes by as
much as 5-fold. We have also shown that newly synthesized class I molecules
bind the TAP transporter (via another protein termed tapasin), an interaction
that
boosts the efficiency of peptide capture by class I.
The functions of tapasin
remain enigmatic. Tapasin clearly is required to increase the efficiency
of peptide capture but it remains an open question as to whether this
is due to its role in bridging class I binding to TAP or to some other
functions. Possibilities include maintaining class I in a peptide receptive
state or acting as a "peptide editor", promoting the exchange of low affinity
peptides for those that bind more tightly.
ERp57 is known to
catalyze disulfide bond formation and isomerization and in fact we have
recently shown that ERp57 enhances the rate of disulfide bond formation
in the class I H chain by 5- to 7-fold.
The Bap31 protein
has been shown by others to promote the transport of some proteins out
of the ER to the Golgi complex. Using RNA interference to reduce Bap31
levels we were able to demonstrate that
Bap31enhances the rate of class I export out of the ER and that Bap31
is required for class I molecules to cluster at ER "exit sites".
It is likely that Bap31 serves as a "cargo receptor" for class
I molecules.
Current research efforts
- Determine how tapasin
increases the efficiency of peptide loading. We are examining whether
tapasin has chaperone functions that may increase H chain folding or
subunit assembly. We are also testing whether tapasin promotes exchange
of low affinity peptides for higher affinity peptides, or whether it
stabilizes "empty" class I molecules in a peptide-receptive state.
- Determine the functions
of ERp57 in class I biogenesis. We are testing the effect of ERp57 on
H chain disulfide bond formation and on the overall efficiency of peptide
loading.
- Ascertain how fully
peptide loaded class I molecules are exported from the ER. Are there
specific receptors that bind to assembled class I and mediate their
export to the Golgi apparatus? A number of candidate receptors are being
examined and efforts are underway to interfere with their functions.
- Using a proteomic
approach, we are searching for additional proteins that participate
in the biogenesis of class I molecules. Candidate proteins are being
characterized and their functions in class I biogenesis assessed.
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