Intermediate-type 20 S Proteasomes in HeLa Cells: “Asymmetric” Subunit Composition, Diversity and Adaptation

The 20 S proteasomes are cylinder-shaped heteromeric dimers with a subunit configuration of α7, β7, β7, α7. Replacement of the three active site-containing standard β-subunits (β1, β2, β5) by immuno-β-subunits (β1i, β2i, β5i) results in formation of 20 S immuno-proteasomes, while only partial replacement leads to intermediate-type proteasomes. Synthesis of immuno-subunits can be induced by interferon-γ, which causes a complete transformation of three subtypes of standard proteasomes into three subtypes of intermediate-type proteasomes in HeLa cells, a process that results in a change in the proteolytic activities of the enzymes. HeLa cells producing the proteasome β1-subunit tagged with the Fc region-binding ZZ domain of protein A were grown in the presence of interferon-γ. From these cells, we have purified 20 S proteasomes by using IgG-affinity resin and analysed them by 2D PAGE. Our study showed that subunit replacement can be confined to one half of the proteasome cylinder, resulting in the formation of intermediate-type proteasomes with ”asymmetric” subunit composition. Analysis of proteasomes purified from the cytoplasm, nucleoplasm, and microsomes of HeLa S3 cells reveals that all three compartments are furnished with intermediate-type proteasomes of different subtype and subunit composition, exhibiting different specific proteolytic activities.