Cryo-EM of mammalian PA28αβ-iCP immunoproteasome reveals a distinct mechanism of proteasome activation by PA28αβ

Abstract

The proteasome activator PA28αβ affects MHC class-I antigen presentation by associating with immunoproteasome core particles (iCPs). However, due to the lack of a mammalian PA28αβ-iCP structure, how PA28αβ regulates proteasome remains elusive. Here we present the complete architectures of the mammalian PA28-iCP immunoproteasome and free iCP at near atomic-resolution by cryo-EM, and determined the spatial arrangement between PA28αβ and iCP through XL-MS. Our structures revealed a slight leaning of PA28αβ towards the 3-4 side of iCP, disturbing the allosteric network of the gate-keeper 3/4 subunits, resulting in a partial open iCP gate. We found that the binding and activation mechanism of iCP by PA28αβ is distinct from those of constitutive CP by the homoheptameric TbPA26 or PfPA28. Our study sheds lights on the mechanism of enzymatic activity stimulation of immunoproteasome and suggests that PA28αβ-iCP has experienced profound remodeling during evolution to achieve its current level of function in immune response.

Conformational maps of human 20S proteasomes reveal PA28- and immuno-dependent inter-ring crosstalks

“…Different sets of α Nter were destabilized depending on the PA28/20S pair (in red Fig. 8c), suggesting a gradual interaction-specific opening of the central pore, in-line with a recent cryo-EM structure that showed a partial opening of the bovine i20S gate in complex with human PA28αβ 65 . The β2 C-ter extending towards β3 was destabilized in all the conditions, except std20S/PA28γ.…”

Allosteric regulation of the proteasome’s catalytic sites by the proteasome activator PA28γ /REGγ

“…Interestingly a mechanism for stimulating unfolded-protein degradation has not been worked out but is expected to be similar. Recently, two cryo-EM studies substantiated similar mechanisms of proteasomal regulation for PA28αβ and the immunoproteasome, and the Pf PA28-20S complex, indicating that PA26, PA28αβ, and Pf PA28 use similar mechanisms to regulate proteasome function 13,26 . In contrast, since its characterization in the 1990s, PA28γ’s effect on the proteasome has been under contention.…”