Substrate-induced protein stabilization reveals a predominant contribution from mature proteins to peptides presented on MHC class I.

The origin of the MHC class I-presented peptides are thought to be primarily from newly synthesized but defective proteins, termed defective ribosomal products. Most of the data supporting this concept come from studies in which inhibitors of protein synthesis were found to rapidly block Ag presentation even when cells contained a pool of mature proteins. However, these data only indirectly address the origin of presented peptides, and in most studies, the contribution of mature functional proteins to the class I peptide pool has not been directly quantified. In this report, we address the efficiency and contribution of mature proteins using a tetracycline-inducible system to express Ags that are conditionally stabilized upon ligand binding. This system circumvents the use of general inhibitors of protein synthesis to control Ag expression. Moreover, by controlling Ag stabilization, we could investigate whether the degradation of mature Ags contributed to Ag presentation at early and/or late time points. We show that mature proteins are the major contributor of peptides presented on class I for two distinct antigenic constructs. Furthermore, our data show that the protein synthesis inhibitors used previously to test the contribution of defective proteins actually block Ag presentation in ways that are independent from blocking Ag synthesis. These data suggest that for the constructs we have analyzed, mature functional proteins, rather than defective ribosomal products, are the predominant source of MHC class I-presented peptides.