0022-1767/91/14710-3306$02.00/0 THE JOURNAL OF ~MMUNOLOCY Copyright 0 1991 by The Amerlcan Association of Immunologists Voi. 147.3306-3313, No. 10. November 15. 1991 Prlnted In U.S.A. PROCESSING OF AN ENDOGENOUS PROTEIN CAN GENERATE MHC CLASS THOSE DERIVED FROM EXOGENOUS ANTIGEN 11-RESTRICTED T CELL DETERMINANTS DISTINCT FROM JOSE MORENO? DARIO A. A. VIGNALI,* FARSIN NADIMI,* SERGE FUCHS,~ LUCIANO ADORINI,~ AND GUNTER J. HAMMERLING" From the *Institute for Immunology and Genetics, German Cancer Research Center [DKFZ), lrn Neuenhelmer Feld 280, 0-6900 Heidelberg, Germany; and 'Sandoz Preclinical Research, CH-4002 Basel, Switzerland Class I1 MHC molecules on the surface of an APC present immunogenic peptides derived mainly from exogenousproteins to CD4+ T cells. During its trans- port to the cell surface, class XI molecules intersect the endocytic pathway where they acquire peptides derived from endocytosed proteins. However, class 11-restricted presentation of endogenously derived peptides can also occur. The current studies were undertaken to examine the ability of different types of APC to generate and present four different T cell determinants derived from an endogenous, nonse- creted, truncated form of hen-egg white lysozyme (HEL[1-80]-Kk). This was compared with the ability of these APC to generate the same determinants from exogenous HEL. All the peptides derived from endogenous HEL[ 1-801-Kk tested, were presented by B cells to HEL-specific T cell hybridomas with an efficiency similar to presentation of the same deter- minants from exogenous HEL. In contrast, an 1-A"- bearing rat fibroblast was unable to generate the HEL peptide 25-43 from exogenous HEL, but could efficiently produce it from endogenous HEL[ 1-801- K". The results indicate first, that peptides derived from an endogenous Ag can be presented by MHC class I1 molecules with an efficiency comparable to that of the presentation of the exogenous Ag. Sec- ond, that Ag-presenting B cells can generate the same repertoire of antigenic peptides from endoge- nous Ag as those generated from the exogenous protein. And third, that in contrast to B cells, cer- tain "nonprofessional" APC can generate, from an endogenous protein, T cell determinants distinct from those generated after endocytosis of the ex- ogenous protein. These results suggest that proc- essing of exogenous and endogenous Ag by different APC take place in different intracellular compart- ments. Unlike Ig, the TCR recognizes protein Ag a s peptides associated to MHC molecules on the surface of an APC (1). The MHC class I molecules generally bind and present peptides derived from proteins synthesized within the Received for publication April 24, 1991. Accepted for publication August 22. 1991. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked aduertlsernent in accordance with 18 U.S.C. Section 1734 solely to indi- cate this fact. Address correspondence and reprint requests to Dr. Jose Moreno. Department of Rheumatology Hospital de Especialidades, Centro Medico Nacional Siglo XXl, IMSS, Av. Cuauhtemoc No. 330. Col. Doctores. 06720 Mtxico, D. F., Mexico. cell (2) and are recognized by CD8+ T cells (3), whereas class I1 MHC molecules mainly bind peptides derived from endocytosed proteins (4) and present them to CD4+ T lymphocytes. The preferential association of endoge- nously or exogenously derived peptides with class I or I1 molecules, respectively, appears to be dictated by their different intracellular routes of transport to the cell sur- face. Thus, class I molecules are exported directly from the Golgi apparatus to the plasma membrane through the constitutive secretory pathway, whereas newly synthe- sized class I1 molecules, before reaching the cell surface, pass through an endocytic compartment in which they meet peptides derived from endocytosed proteins (5-7). Acidification of endosomes or lysosomes is a necessary step for antigen processing and presentation by class I1 (8-101 but not class I MHC molecules (2). However, recognition of endogenously derived peptides in the context of class I1 molecules has now been exten- sively documented [ 1 1 - 17), but it is not universal (2, 18. 19). Processing of endogenous proteins for presentation by class I1 can occur in the endocytic pathway (1 1). Yet, some endogenous proteins appear to be processed in an intracellular compartment that is independent of acidi- fication (14, 16). The characteristics of an endogenous protein needed for presentation by class I1 molecules are not well understood. A recent study suggests that only proteins that are translocated into the ER2 can be pre- sented by class I1 molecules (17). Nevertheless, other studies have found class 11-restricted presentation of pep- tides derived from purely cytosolic proteins (14, 20, 21). Processing of a protein Ag by certain types ofAPC or in different intracellular compartments could generate dis- tinct T cell determinants. Supporting the potential im- portance of this is the variability in the intracellular distribution of proteases in different cell types (22), as well as the observation that certain determinants of a given protein may be the result of different proteases (23, 24). To further pursue these issues, a constructencoding a truncated, nonsecreted form of hen egg-white lysozyme (HEL[ 1 -80]-Kk) was generated and transfected into two differenttypes of APC. Using these transfectants we investigated whether the first 80 amino acids of exoge- nous HEL and endogenous HEL( 1 -801-Kk could generate the same repertoire of immunogenic peptides. 'Abbreviations used in this paper: ER, endoplasmic reticulum: HEL. hen-egg white lysozyme: HEL[1-801-Kk, chimeric protein, product of the fusion of HEL and H-2Kk genes: GaRIg (GaMIg).goat anti-rat (mouse) Ig: MIIC. class 11-related endocytic compartment: neo, neomycin-resistance gene: li. class Il-associated invariant chain: TM. transmembrane domain. 3306