Plasticity in eucaryotic 20S proteasome ring assembly revealed by a subunit deletion in yeast

Abstract

The 20S proteasome is made up of four stacked heptameric rings, which in eucaryotes assemble from 14 different but related subunits. The rules governing subunit assembly and placement are not understood. We show that a different kind of proteasome forms in yeast when the Pre9/α3 subunit is deleted. Purified pre9A proteasomes show a two-fold enrichment for the Pre6/α4 subunit, consistent with the presence of an extra copy of Pre6 in each outer ring. Based on disulfide engineering and structure-guided suppressor analyses, Pre6 takes the position normally occupied by Pre9, a substitution that depends on a network of intersubunit salt bridges. When Arabidopsis PAD1/α4 is expressed in yeast, it complements not only pre6Δ but also pre6Δ pre9δ mutants; therefore, the plant α4 subunit also can occupy multiple positions in a functional yeast proteasome. Importantly, biogenesis of proteasomes is delayed at an early stage in pre9Δ cells, suggesting an advantage for Pre9 over Pre6 incorporation at the α3 position that facilitates correct assembly.