ABSTRACT

The antigen presentation pathway yields peptide-MHC class I complexes on the antigen presenting cell (APC) surface for recognition by appropriate T-cells. Expression of the peptide-MHC complex on APC surface is preceded by several steps that include the generation of peptide fragments in the cytoplasm and their assembly with MHC molecules in the endoplasmic reticulum. It is now clear that MHC binding to optimally processed peptides in the endoplasmic reticulum is obligatory for their stable expression on the cell surface. However, whether a similar obligatory relationship exists between generation of processed peptides and their expression as peptide-MHC on APC surface is not known. Here, we addressed this question by analyzing the processing of ovalbumin (aa257-264, SL8) or influenza nucleoprotein (aa366-374, AM9) analogs. We examined the generation of naturally processed peptides using precursors that did, or did not, contain residues flanking the optimal MHC-binding peptides. By characterizing the peptides generated from these precursors by T-cell stimulation assays and by high performance liquid chromatography analysis, we established that intracellular assembly of peptide-MHC complexes and their expression on the cell surface can occur with peptides that lack flanking residues. The presentation of these endogenously synthesized perfect fit peptides demonstrates that the cleavage of precursor polypeptides is an independent step in the antigen presentation pathway.

FOOTNOTES

*

This research was supported by National Institutes of Health Grant AI-26604 and by a grant from the UC Tobacco Related Disease Research Grant (to N. S.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked ``advertisement'' in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

§

Recipient of a pre-doctoral fellowship from the Howard Hughes Medical Institute.

¶

Recipient of an University of California System-wide Biotechnology Research and Education Program Training Grant.

**

To whom correspondence should be addressed: Dept. of Molecular Cell Biology LSA 421, University of California, Berkeley, CA 94720-3200. Tel.: 510-643-9197; Fax: 510-643-9230.

()

The abbreviations used are: MHC, major histocompatibility complex; APC, antigen presenting cell; ER, endoplasmic reticulum; PBS, phosphate-buffered saline; HPLC, high performance liquid chromatography.

()

P. Scott, unpublished results.

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S. Goth and N. Shastri, unpublished data.

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