Interaction with the Assembly Chaperone Ump1 Promotes Incorporation of the β7 Subunit into Half-Proteasome Precursor Complexes Driving Their Dimerization.

Jessica Zimmermann,R Jürgen Dohmen,Paula C Ramos

doi:10.3390/biom12020253

Abstract

Biogenesis of the eukaryotic 20S proteasome core particle (PC) is a complex process assisted by specific chaperones absent from the active complex. The first identified chaperone, Ump1, was found in a precursor complex (PC) called 15S PC. Yeast cells lacking Ump1 display strong defects in the autocatalytic processing of β subunits, and consequently have lower proteolytic activity. Here, we dissect an important interaction of Ump1 with the β7 subunit that is critical for proteasome biogenesis. Functional domains of Ump1 and the interacting proteasome subunit β7 were mapped, and the functional consequences of their deletion or mutation were analyzed. Cells in which the first sixteen Ump1 residues were deleted display growth phenotypes similar to ump1∆, but massively accumulate 15S PC and distinct proteasome intermediate complexes containing the truncated protein. The viability of these cells depends on the transcription factor Rpn4. Remarkably, β7 subunit overexpression re-established viability in the absence of Rpn4. We show that an N-terminal domain of Ump1 and the propeptide of β7 promote direct interaction of the two polypeptides in vitro. This interaction is of critical importance for the recruitment of β7 precursor during proteasome assembly, a step that drives dimerization of 15S PCs and the formation of 20S CPs.

Highlights

Intracellular protein degradation is essential for protein quality control, as well as the regulation of cellular processes
To learn more about how the truncations affect the function of Ump1, we asked if the different versions would be competent to form 15S precursor complex (PC)
All of the tested truncated versions 17–148Ump1, 56–148Ump1, and 82–148Ump1 were detected in 15S PCs as well, indicating that residues 82–148 are sufficient for the incorporation of Ump1 into these complexes

Summary

Introduction

Intracellular protein degradation is essential for protein quality control, as well as the regulation of cellular processes. In contrast to the two dimeric Pba chaperones that are recycled during the biogenesis process, Ump is degraded by the native 20S proteasome upon completion of its assembly [5,6]. Deletion of the UMP1 gene produces cells impaired in proteasomal activity with strong defects in autocatalytic processing of the active β-subunits [5]. A bulkier extension such as a GFP module hocked to the N terminus of Ump blocks dimerization so dramatically that the pathway fails to produce fully assembled proteasomes, resulting in the severe inhibition of cell growth [6]. Yeast cells are able to form proteasomes in the total absence of the chaperone Ump, the growth defects and proteasome impairment indicate that PC biogenesis is inefficient and error-prone [5]

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