title: Cdc48-independent proteasomal degradation coincides with a reduced need for ubiquitylation.
abstract: Ubiquitin fusion degradation (UFD) substrates are delivered at the proteasome by a handover mechanism involving the ubiquitin-selective chaperone Cdc48 and the ubiquitin shuttle factor Rad23. Here, we show that introduction of a 20 amino acid peptide extension not only rendered degradation independent of Cdc48, in line with the model that this chaperone is involved in early unfolding events of tightly folded substrates, but at the same time relieved the need for efficient polyubiquitylation and the ubiquitin shuttle factor Rad23. Removal of the ubiquitylation sites in the N-terminal UFD signal made the degradation of this substrate strictly dependent on the peptide extension and also on Cdc48 and, importantly the presence of a functional ubiquitylation machinery. This suggests that the extension in the absence of N-terminal ubiquitylation sites is not properly positioned to engage the unfoldase machinery of the proteasome. Thus the need for efficient ubiquitylation and Cdc48 in facilitating proteasomal degradation are tightly linked but can be bypassed in the context of UFD substrates by the introduction of an unstructured extension. Our data suggest that polyubiquitin-binding complexes acting upstream of the proteasome, rather than the proteasome itself, can be primary determinants for the level of ubiquitylation required for protein degradation.
authors.affiliation: Department of Cell and Molecular Biology, Karolinska Institutet, S-17177 Stockholm, Sweden.
authors.forename: Daniela
authors.initials: D
authors.lastname: Gödderz
authors.affiliation: Department of Cell and Molecular Biology, Karolinska Institutet, S-17177 Stockholm, Sweden.
authors.forename: Christian
authors.initials: C
authors.lastname: Heinen
authors.affiliation: Department of Cell and Molecular Biology, Karolinska Institutet, S-17177 Stockholm, Sweden.
authors.forename: Francesco P
authors.initials: FP
authors.lastname: Marchese
authors.affiliation: Department of Cell and Molecular Biology, Karolinska Institutet, S-17177 Stockholm, Sweden.
authors.forename: Tilman
authors.initials: T
authors.lastname: Kurz
authors.affiliation: Department of Cell and Molecular Biology, Karolinska Institutet, S-17177 Stockholm, Sweden.
authors.forename: Klàra
authors.initials: K
authors.lastname: Acs
authors.affiliation: Department of Cell and Molecular Biology, Karolinska Institutet, S-17177 Stockholm, Sweden.
authors.forename: Nico P
authors.initials: NP
authors.lastname: Dantuma
mesh_terms: D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D059748:Proteolysis; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity; D025801:Ubiquitin / Q000378:metabolism; D054875:Ubiquitination; D000074405:Valosin Containing Protein
chemical_list: D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C490846:RAD23 protein, S cerevisiae; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin; D049452:Green Fluorescent Proteins; D046988:Proteasome Endopeptidase Complex; D000251:Adenosine Triphosphatases; C000614177:CDC48 protein, S cerevisiae; D000074405:Valosin Containing Protein