Influence of the tapasin C terminus on the assembly of MHC class I allotypes.

Several endoplasmic reticulum proteins, including tapasin, play an important role in major histocompatibility complex (MHC) class I assembly. In this study, we assessed the influence of the tapasin cytoplasmic tail on three mouse MHC class I allotypes (H2-K(b), -K(d), and -L(d)) and demonstrated that the expression of truncated mouse tapasin in mouse cells resulted in very low K(b), K(d), and L(d) surface expression. The surface expression of K(d) also could not be rescued by human soluble tapasin, suggesting that the surface expression phenotype of the mouse MHC class I molecules in the presence of soluble tapasin was not due to mouse/human differences in tapasin.

Amyloid precursor-like protein 2 increases the endocytosis, instability, and turnover of the H2-K(d) MHC class I molecule.

The defense against the invasion of viruses and tumors relies on the presentation of viral and tumor-derived peptides to CTL by cell surface MHC class I molecules. Previously, we showed that the ubiquitously expressed protein amyloid precursor-like protein 2 (APLP2) associates with the folded form of the MHC class I molecule K(d). In the current study, APLP2 was found to associate with folded K(d) molecules following their endocytosis and to increase the amount of endocytosed K(d).

Effect of linkage of transduction domain sequences to a lymphoma idiotype DNA vaccine on vaccine effectiveness.

Patient idiotype-specific vaccines for treatment of non-Hodgkin's lymphoma have shown promise in clinical trials, encouraging efforts to enhance the effectiveness of idiotype vaccines further. It has previously been found that for some other types of experimental vaccines, the addition of transduction domains has improved vaccine immunogenicity. Transduction domains are short amino acid sequences that are capable of increasing transport through cellular membranes.

A charged amino acid residue in the transmembrane/cytoplasmic region of tapasin influences MHC class I assembly and maturation.

Tapasin influences the quantity and quality of MHC/peptide complexes at the cell surface; however, little is understood about the structural features that underlie its effects. Because tapasin, MHC class I, and TAP are transmembrane proteins, the tapasin transmembrane/cytoplasmic region has the potential to affect interactions at the endoplasmic reticulum membrane. In this study, we have assessed the influence of a conserved lysine at position 408, which lies in the tapasin transmembrane/cytoplasmic domain.

Tapasin decreases immune responsiveness to a model tumor antigen.

The T-cell response against cancer is dependent on the cell surface presentation of tumor-associated or tumor-specific peptides by major histocompatibility complex (MHC) class I molecules. We found that tapasin, a chaperone protein that normally assists in the assembly of MHC class I molecules, is undetectable in an unstimulated pancreatic tumor cell line, Panc02, and only very weakly expressed after gamma-interferon stimulation. Transfection of tapasin into the Panc02 cells did not quantitatively increase MHC class I surface expression or detectably affect MHC class I association with peptide and beta(2)-microglubulin (beta(2)m).

The Ig-like domain of tapasin influences intermolecular interactions.

Presentation of antigenic peptides to T lymphocytes by MHC class I molecules is regulated by events involving multiple endoplasmic reticulum proteins, including tapasin. By studying the effects of substitutions in the tapasin Ig-like domain, we demonstrated that H-2L(d)/tapasin association can be segregated from reconstitution of folded L(d) surface expression. This finding suggests that peptide acquisition by L(d) is influenced by tapasin functions that are independent of L(d) binding.

The amyloid precursor-like protein 2 and the adenoviral E3/19K protein both bind to a conformational site on H-2Kd and regulate H-2Kd expression.

A protein of unknown physiological function, called amyloid precursor-like protein 2 (APLP2), forms an association with the murine class I molecule K(d) that is up-regulated by the presence of the adenoviral protein E3/19K. We have extended these findings to show that APLP2 and E3/19K associate preferentially with folded K(d) and not with the open form. APLP2 was detectable at the cell surface, but its surface expression was not up-regulated by the concurrent expression of K(d).

HLA class I polymorphism has a dual impact on ligand binding and chaperone interaction.

This article will describe coordinated analyses of how amino acid substitutions in the HLA class I antigen binding groove modify chaperone interaction and peptide ligand presentation. By parallel testing of ligand presentation and chaperone interaction with a series of natural HLA-B subtypes, this study has discovered that position 116 of the HLA-B15 class I heavy chain is pivotal in both peptide selection and control of interaction between the assembly complex and the class I heavy chain. Correlated with these qualitative differences in peptide selection and chaperone association are quantitative differences in the expression levels of the HLA molecules at the cell surface.

The interface between tapasin and MHC class I: identification of amino acid residues in both proteins that influence their interaction.

Prior to the binding of antigenic peptide, a complex of chaperone proteins associates with the Major Histocompatibility Complex (MHC) class I heavy chain/beta2m heterodimer. Although each domain of the MHC class I heavy chain contains amino acid residues that influence chaperone binding, there are several pieces of evidence that point to an interaction between the MHC class I alpha/2/alpha3 domains and tapasin. In regard to the site on tapasin involved in the tapasin/MHC interface, we have found that a particular region of tapasin (containing amino acid residues 334-342) is necessary for the binding of tapasin to the MHC class I heavy chain.

Disparate binding of chaperone proteins by HLA-A subtypes.

We examined chaperone association with subtypes of HLA-A68 differing at positions 116 and/or 70, and analyzed the surface expression of each A68 subtype. Our findings with A68 indicate that certain subtypes have inefficient association with the assembly complex and correspondingly high surface expression, dependent on the character of position 116. Specifically, poor association of A68 subtypes with the transporter associated with antigen processing correlated with a comparatively high level of W6/32(+) forms at the cell surface.