Antigen processing and presentation

0

INSTITUT

Res. Immunol.

PASTEUR/ELSEVIER

Paris 1995

1995 ENII SUMMARY

1995, 146, 397-413

CONFERENCE

OF SESSIONS

Antigen processing and presentation Chairperson

: J. Neefjes

The field of antigen processing and presentation is progressing very rapidly. Various presentations have basically covered this field. The first half of the session was dedicated to different steps in antigen presentation by class I molecules, while the second half of the session discussed antigen presentation by class II molecules and some functional consequences of antigen interaction with antibodies or surface receptors. Finally, the identification of minor histocompatibility antigens was discussed. It has been recognized for more than a decade that MHC class I molecules present fragments of cytosolic and nuclear antigens. Half a decade ago, it was established that these fragments were usually of defined length, of = 9 amido acids. Presentation of such a fragment should be preceded by a number of events: the antigen should first be degraded, and the degradation fragment should be translocated from the cytosol into the ER lumen and associate with MHC class I molecules. K. Rock (Boston) discussed his studies on cytosolic degradation of antigen. He first introduced the current view on cytosolic degrada-

(Amsterdam)

tion. Proteins destined for degradation are usually tagged first by a poly-ubiquitin tree. This requires the successive action of a number of enzymes, named El, E2 and E3, respectively. The protein is then degraded by a multi-enzyme complex, the 20s and the 26s proteasome. Using hamster cells with a ts-mutation in El (thus affecting ubiquitin addition), Rock showed that presentation of soluble ovalbumin was dependent on El action. Depending on the first amino acid, proteins can be more or less susceptible to ubiquitin-dependent degradation (the so called N-end rule). Indeed, the half-life of a protein may affect presentation by class I molecules. Rock used a number of inhibitors for proteasomes to show that they inhibited peptide generation and assembly of class I molecules. These inhibitors are not entirely specific, but by using inhibitors with different specificities, involvement of proteasomes in antigen degradation is made likely. J. Trowsdale (London) described the isolation of two proteasome subunit genes (Imp2 and Imp7) located in close proximity to the TAP genes in the MHC locus. These genes are upregulated by y-IFN treatment and studies with mice with

Received August 30, 1995. For reprint requests of the Summary of Sessions, strasse 487, Postgach, CH-4005 Basel, Switzerland.

please contact

Dr. A. Lanzavecchia,

Base1 Institute

for Immunology,

Grenzacher-

398

SUMMARY

inactivated Imp2 or Imp7 genes showed that they influence the generation and presentation of antigen. All these data indicate that proteasomes can play a role in antigen generation but do not exclude the involvement of other pathways in antigen degradation. The next step is translocation of a peptide over the ER membrane. Studies on mutant cell lines had already indicated the involvement of a heterodimer of two multimembrane spanning proteins, TAP1 and TAP2, in this process. J. Neefjes (Amsterdam) showed the size and sequence specificity of TAP of three species. TAP has a broad sequence specificity and prefers peptides of 8-13 amino acids in length, although longer peptides can be translocated. Thus, the peptide specificity of TAP matches that of class I molecules. The fate of peptides that fail to bind to class I molecules was followed. Although some degradation of peptides in the ER could be detected, most peptides were rapidly released into the cytosol where they were either degraded or recycled back into the ER lumen. Nijenhuis (Heidelberg) directly visualized peptide binding to TAP using photocross-linker-containing peptides. These peptides are now used to define the peptide-contact areas of TAP. The majority of class I-presented peptides are from an endogenous source. Watts (Dundee) showed that exogenous antigen can also enter the class I pathway by macropinocytosis. Macropinocysis can be induced by massive internalization of the EGT-receptor or by stimulation of macrophages. Soluble antigen can then enter the cytosol. Possibly this pathway permits new immunization strategies using intact antigens. MI-K class II molecules present fragments of endocytosed antigen. These fragments are generated deep in the endocytic pathway where class II molecules associate with fragments prior to cell surface exposure. Analysis of mutant cell lines indicated that certain proteins, again encoded in the MHC locus, should be involved in efficient loading of MHC class II molecules. J. Trowsdale (London) discussed the identification of the proteins involved, HLA-DMA and -DMB, that together form a heterodimer with presumably a

OF SESSIONS

structure homologous to class I and II molecules. HLA-DM is located in a lysosomal-like structure, previously named MIIC, and arrives there because the HLA-DMB chain contains a lysosoma1 targetting signal. The exact action of HLADM is unclear. Class II molecules are targetted into the endocytic pathway by the associated invariant chain or li that exists in different spliced forms. Fineschi (Chicago) reported an effect of degradation of the p41 form that exclusively results in the formation of a 12 kDa N-terminal fragment. However, the degradation of the p31 form seems to be affected by the p41 form. How this should explain the superior antigen presentation capacity of p41 is unknown. Although li prevents premature binding of peptides in the ER, endogenous antigen presentation is still observed. Oxenius (Zurich) studied this process by following the presentation of the LCMV-nucleoprotein (NP) and glycoprotein (GP). In contrast to GP, NP presentation by class II molecules was blocked with endosomal/lysosomal inhibitors. However, neither GP nor NP require li or TAP. How NP fragments enter endosomes is unclear. For presentation of GP, it is anticipated that GP is first inserted into the ER followed by degradation, and that class II molecules then associate with peptide in the ER. How the generation and presentation of antigen may be modulated is unclear. Both Watts (Dundee) and Barnaba (Rome) presented different examples showing how receptor/antigen interactions may influence the degradation of the antigen and thereby hide or expose certain presentable epitopes. Watts showed this for uptake of antigen by surface immunoglobulin on B cells. Bamaba showed that epitopes of CD4 molecules that have not been presented during T-cell selection may be presented when HIV gp 120 increases internalization and processing of CD4 such that hidden CD4 epitopes can be unveiled. This may then result in an autoimmune-like T-cell response resulting in elimination of the CD4+ T cells. Also, changes in host protein synthesis may result in exposure of cryptic peptides from self proteins, and Bamaba showed that CTL against vinculin can be generated against HIVinfected PBL. Exposure of cryptic peptides may be of great relevance for autoimmune responses

IMMUNE

(see also Lanzavecchia, (1995)).

SYSTEM PHYSIOLOGY

J.E.M.

181,

1945

Finally, two presentations showed the identification of peptides that had long been searched for. Den Haan (Leiden) characterized the minor histocompatibility antigen HA-2 presented by HLA-A2 using state of the art microcapillary HPLUelectron spray ionization tandem mass spectrometry. A peptide was found that was recognized by specific CTL and is possibly derived from an as yet unknown myosin family member. This information is to be used for better typing of transplants. Ehrmann (London) identified the male-specific antigen HY using a more genetic approach. A gene (SMCY)

Lymphocyte Chairperson

: B. Malissen

T-cell activation is controlled by the antigen receptor TCR/CD3 complex. The cytoplasmic segments of the various CD3 chains are responsible for coupling the TCR to intracellular signalling pathways. Their signal transduction capability has been attributed to the presence of a recurrent functional domain referred to as the immunoreceptor tyrosine-based activation motif (ITAM). Once phosphorylated, via protein tyrosine kinases (PTK) of the src family (Ick orfyn), the two YXXL/I sequences present within a given ITAM are capable of being recognized in a concerted mode by the tandem SH2 domains present in the ZAP-70 and syk PTKs. At least two PTK-controlled signalling pathways originate from the TCR. One involves the guanine nucleotide binding proteins p2lras and the other is mediated by phospholipase C (PLC). PLC regulates the hydrolysis of inositol phospholipids and thereby allows the TCR to regulate intracellular

AND PATHOLOGY

399

is identified on the short arm of murine Y chromosomes that is recognized by HY-specific CTL. Poster presentations also covered many different topics including correct predictions of presented peptides, peptide competition by different class I molecules, introduction of soluble exogenous antigen into the class I pathway, invariant chain interactions with class II molecules, the role of HLA-DM in antigen presentation by class II molecules as well as the invariant chain and the function of non-classical MHC molecules. This shows that antigen processing and presentation is not only a rapidly progressing but also a lively field.

activation (CIML,

Marseille)

calcium metabolism and the serine/threonine kinase family of PKC isozymes. The details of TCR regulation of p2lras are poorly understood but probably involve TCR regulation of p2lras GTPase activating proteins and p2lras guanine nucleotide exchange proteins. The recent advances in understanding how p21 ras couples the TCR to the T-cell signalling cascade have been reviewed by J. Nunes (London). One TCR/p2lras link is via the guanine nucleotide exchange proteins SOS which is regulated by the adapter protein Grb-USemS. Grb-2 is composed of one SH2 domain and two SH3 domains. The SH3 domains of Grb2 bind to the carboxy terminal proline-rich domain of SOS and the interactions of the Grbr2 SH2 domain with tyrosinephosphorylated molecules apparently recruits SOS to the cell membrane. She and a membranebound tyrosine phosphotrotein of 36 kDa are tyrosine phosphorylated in TCR-activated T cells