Abstract
BackgroundHistone H2B deubiquitination is performed by numerous deubiquitinases in eukaryotic cells including Ubp8, the catalytic subunit of the tetrameric deubiquitination module (DUBm: Ubp8; Sus1; Sgf11; Sgf73) of the Spt-Ada-Gcn5 acetyltransferase (SAGA). Ubp8 is linked to the rest of SAGA through Sgf73 and is activated by the adaptors Sus1 and Sgf11. It is unknown if DUBm/Ubp8 might also work in a SAGA-independent manner.ResultsHere we report that a tetrameric DUBm is assembled independently of the SAGA–CORE components SPT7, ADA1 and SPT20. In the absence of SPT7, i.e., independent of the SAGA complex, Ubp8 and Sus1 are poorly recruited to SAGA-dependent genes and to chromatin. Notably, cells lacking Spt7 or Ada1, but not Spt20, show lower levels of nuclear Ubp8 than wild-type cells, suggesting a possible role for SAGA–CORE subunits in Ubp8 localization. Last, deletion of SPT7 leads to defects in Ubp8 deubiquitinase activity in in vivo and in vitro assays.ConclusionsCollectively, our studies show that the DUBm tetrameric structure can form without a complete intact SAGA–CORE complex and that it includes full-length Sgf73. However, subunits of this SAGA–CORE influence DUBm association with chromatin, its localization and its activity.
Highlights
Histone H2B deubiquitination is performed by numerous deubiquitinases in eukaryotic cells including Ubp8, the catalytic subunit of the tetrameric deubiquitination module (DUBm: Ubp8; Sus1; Sgf11; Sgf73) of the SptAda-Gcn5 acetyltransferase (SAGA)
A tetrameric DUBm containing full‐length Sgf73 can form without a complete intact SAGA–core” module (CORE) complex The DUBm can be separated from the rest of SAGA by increasing salt concentration or by deleting SAGA subunits such as SGF73 or SPT20 [25,26,27,28]
In the light of these new structural findings, we determined if Spt7 or Ada1 might play a role in DUBm assembly by investigating the ability of the DUBm to assemble in the absence of these subunits of the SAGA–CORE
Summary
Histone H2B deubiquitination is performed by numerous deubiquitinases in eukaryotic cells including Ubp, the catalytic subunit of the tetrameric deubiquitination module (DUBm: Ubp; Sus; Sgf; Sgf73) of the SptAda-Gcn acetyltransferase (SAGA). Two new studies reporting cryo-EM structures significantly increased our knowledge of SAGA stoichiometry and protein–protein interactions [17, 18] They show that most of the proteins from the modules previously known as SPT and TAF form a structural CORE complex involving Spt, Spt, Spt, Ada, Taf, Taf, Taf, Taf, Taf and some residues of Sgf and Ada3 [19]. The octamer-like structure is composed of paired HF proteins linked to each other in the order: an initial HF Taf6–Taf pair, followed by the Taf12–Ada pair, the Taf10–Spt pair, and a final Spt HF pair [19] Those studies further revealed a close connection between the HAT and the DUB activities of SAGA on chromatin
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