title: Proteasome-mediated processing of Def1, a critical step in the cellular response to transcription stress.
abstract: DNA damage triggers polyubiquitylation and degradation of the largest subunit of RNA polymerase II (RNAPII), a "mechanism of last resort" employed during transcription stress. In yeast, this process is dependent on Def1 through a previously unresolved mechanism. Here, we report that Def1 becomes activated through ubiquitylation- and proteasome-dependent processing. Def1 processing results in the removal of a domain promoting cytoplasmic localization, resulting in nuclear accumulation of the clipped protein. Nuclear Def1 then binds RNAPII, utilizing a ubiquitin-binding domain to recruit the Elongin-Cullin E3 ligase complex via a ubiquitin-homology domain in the Ela1 protein. This facilitates polyubiquitylation of Rpb1, triggering its proteasome-mediated degradation. Together, these results outline the multistep mechanism of Rpb1 polyubiquitylation triggered by transcription stress and uncover the key role played by Def1 as a facilitator of Elongin-Cullin ubiquitin ligase function.
authors.affiliation: Mechanisms of Transcription Laboratory, Clare Hall Laboratories, Cancer Research UK London Research Institute, South Mimms EN6 3LD, UK.
authors.forename: Marcus D
authors.initials: MD
authors.lastname: Wilson
authors.affiliation: Mechanisms of Transcription Laboratory, Clare Hall Laboratories, Cancer Research UK London Research Institute, South Mimms EN6 3LD, UK.
authors.forename: Michelle
authors.initials: M
authors.lastname: Harreman
authors.affiliation: Mechanisms of Transcription Laboratory, Clare Hall Laboratories, Cancer Research UK London Research Institute, South Mimms EN6 3LD, UK.
authors.forename: Michael
authors.initials: M
authors.lastname: Taschner
authors.affiliation: Mechanisms of Transcription Laboratory, Clare Hall Laboratories, Cancer Research UK London Research Institute, South Mimms EN6 3LD, UK.
authors.forename: James
authors.initials: J
authors.lastname: Reid
authors.affiliation: Mechanisms of Transcription Laboratory, Clare Hall Laboratories, Cancer Research UK London Research Institute, South Mimms EN6 3LD, UK.
authors.forename: Jane
authors.initials: J
authors.lastname: Walker
authors.affiliation: Molecular Biology Programme, Memorial Sloan-Kettering Cancer Center, York Avenue 1275, New York, NY 10021, USA.
authors.forename: Hediye
authors.initials: H
authors.lastname: Erdjument-Bromage
authors.affiliation: Molecular Biology Programme, Memorial Sloan-Kettering Cancer Center, York Avenue 1275, New York, NY 10021, USA.
authors.forename: Paul
authors.initials: P
authors.lastname: Tempst
authors.affiliation: Mechanisms of Transcription Laboratory, Clare Hall Laboratories, Cancer Research UK London Research Institute, South Mimms EN6 3LD, UK. Electronic address: j.svejstrup@cancer.org.uk.
authors.forename: Jesper Q
authors.initials: JQ
authors.lastname: Svejstrup
mesh_terms: D000595:Amino Acid Sequence; D002868:Chromosomal Proteins, Non-Histone / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012319:RNA Polymerase II / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000502:physiology*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*; D016415:Sequence Alignment; D013312:Stress, Physiological; D014158:Transcription, Genetic; D043743:Ubiquitin-Protein Ligase Complexes / Q000378:metabolism
chemical_list: D002868:Chromosomal Proteins, Non-Histone; C451742:DEF1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D043743:Ubiquitin-Protein Ligase Complexes; D012319:RNA Polymerase II; D046988:Proteasome Endopeptidase Complex