title: A SUMO-targeted ubiquitin ligase is involved in the degradation of the nuclear pool of the SUMO E3 ligase Siz1.
abstract: The Slx5/Slx8 heterodimer constitutes a SUMO-targeted ubiquitin ligase (STUbL) with an important role in SUMO-targeted degradation and SUMO-dependent signaling. This STUbL relies on SUMO-interacting motifs in Slx5 to aid in substrate targeting and carboxy-terminal RING domains in both Slx5 and Slx8 for substrate ubiquitylation. In budding yeast cells, Slx5 resides in the nucleus, forms distinct foci, and can associate with double-stranded DNA breaks. However, it remains unclear how STUbLs interact with other proteins and their substrates. To examine the targeting and functions of the Slx5/Slx8 STUbL, we constructed and analyzed truncations of the Slx5 protein. Our structure-function analysis reveals a domain of Slx5 involved in nuclear localization and in the interaction with Slx5, SUMO, Slx8, and a novel interactor, the SUMO E3 ligase Siz1. We further analyzed the functional interaction of Slx5 and Siz1 in vitro and in vivo. We found that a recombinant Siz1 fragment is an in vitro ubiquitylation target of the Slx5/Slx8 STUbL. Furthermore, slx5 cells accumulate phosphorylated and sumoylated adducts of Siz1 in vivo. Specifically, we show that Siz1 can be ubiquitylated in vivo and is degraded in an Slx5-dependent manner when its nuclear egress is prevented in mitosis. In conclusion, our data provide a first look into the STUbL-mediated regulation of a SUMO E3 ligase.
authors.affiliation: Biology Department, The College of William & Mary, Williamsburg, VA 23187.
authors.forename: Jason W
authors.initials: JW
authors.lastname: Westerbeck
authors.forename: Nagesh
authors.initials: N
authors.lastname: Pasupala
authors.forename: Mark
authors.initials: M
authors.lastname: Guillotte
authors.forename: Eva
authors.initials: E
authors.lastname: Szymanski
authors.forename: Brooke C
authors.initials: BC
authors.lastname: Matson
authors.forename: Cecilia
authors.initials: C
authors.lastname: Esteban
authors.forename: Oliver
authors.initials: O
authors.lastname: Kerscher
mesh_terms: D002467:Cell Nucleus / Q000201:enzymology*; D019913:Nuclear Localization Signals; D010766:Phosphorylation; D011485:Protein Binding; D021381:Protein Transport; D059748:Proteolysis; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*; D054875:Ubiquitination
chemical_list: D019913:Nuclear Localization Signals; D029701:Saccharomyces cerevisiae Proteins; D044767:Ubiquitin-Protein Ligases; C436837:Siz1 protein, S cerevisiae; C515573:Slx5 protein, S cerevisiae