Hexameric assembly of the proteasomal ATPases is templated through their C termini

Abstract

The proteasome is a large proteolytic machine that degrades ubiquitin-tagged proteins. Substrates are recognized and unfolded by the regulatory particle (RP) and translocated into a central proteolytic chamber called the core particle (CP) where degradation takes place. The CP associates with the RP at either one or both ends. The RP can be further subdivided into the base and lid. Two studies from Finley and colleagues elucidate the pathway of RP assembly. They report the identification of three molecular chaperones that assist in the assembly of the RP. Assembly of the base proceeds through a complex consisting of five proteins, called BP1, which functions as an intermediate in the process. These studies show that RP assembly is a highly orchestrated process. The proteasome is a large proteolytic machine that degrades ubiquitin-tagged proteins. Substrates are recognized and unfolded by the regulatory particle (RP) and translocated into a central proteolytic chamber, called the core particle (CP), where degradation takes place. Here, and in a complimentary study, the pathway of RP assembly is elucidated, demonstrating that RP assembly is a highly orchestrated process. Substrates of the proteasome are recognized and unfolded by the regulatory particle, and then translocated into the core particle (CP) to be degraded1. A hetero-hexameric ATPase ring, containing subunits Rpt1-6, is situated within the base subassembly of the regulatory particle1. The ATPase ring sits atop the CP, with the Rpt carboxy termini inserted into pockets in the CP2,3,4,5,6. Here we identify a previously unknown function of the Rpt proteins in proteasome biogenesis through deleting the C-terminal residue from each Rpt in the yeast Saccharomyces cerevisiae. Our results indicate that assembly of the hexameric ATPase ring is templated on the CP. We have also identified an apparent intermediate in base assembly, BP1, which contains Rpn1, three Rpts and Hsm3, a chaperone for base assembly. The Rpt proteins with the strongest assembly phenotypes, Rpt4 and Rpt6, were absent from BP1. We propose that Rpt4 and Rpt6 form a nucleating complex to initiate base assembly, and that this complex is subsequently joined by BP1 to complete the Rpt ring. Our studies show that assembly of the proteasome base is a rapid yet highly orchestrated process.