The effect of heat shock on 20S/26S proteasomes.

We have studied the consequences of heat shock on 20S/26S proteasome activity and activation, the proteasomal subunit composition, proteasome assembly, subunit mRNA stability as well as on the intracellular distribution of proteasomes. Our data show that heat shock locks 20S proteasomes in their latent inactive state and impairs further activation of the 26S proteasome by ATP. Proteasome mRNA levels are decreased after heat shock and the assembly of the proteasome complex is inhibited. Heat shock also induces a rapid reorganisation of the cellular distribution of the proteasome which appears to be connected with proteasome activity and the change of the cellular architecture after heat shock.

The cleavage preference of the proteasome governs the yield of antigenic peptides.

Proteasomes degrade endogenous proteins in the cytosol. The potential contribution of the proteasome to the effect of flanking sequences on the presentation of an antigenic epitope presented by the major histocompatibility complex class I allele Ld was studied. Peptides generated in cells from minigenes coding for peptides of 17- and 19-amino acid length were compared with the in vitro 20S proteasome degradation products of the respective synthetic peptides. The quality of generated peptides was independent of ubiquitination. In vivo and in vitro processing products were indistinguishable with respect to peptide size and abundance. Altering the neighboring sequence substantially improved the yield of the final antigenic nonapeptide by 20S proteasome cleavage. These results suggest that, in addition to the presence of major histocompatibility complex class I allelic motifs, the cleavage preference of the proteasome can define the antigenic potential of a protein.