title: Higher-order oligomerization promotes localization of SPOP to liquid nuclear speckles.
abstract: Membrane-less organelles in cells are large, dynamic protein/protein or protein/RNA assemblies that have been reported in some cases to have liquid droplet properties. However, the molecular interactions underlying the recruitment of components are not well understood. Herein, we study how the ability to form higher-order assemblies influences the recruitment of the speckle-type POZ protein (SPOP) to nuclear speckles. SPOP, a cullin-3-RING ubiquitin ligase (CRL3) substrate adaptor, self-associates into higher-order oligomers; that is, the number of monomers in an oligomer is broadly distributed and can be large. While wild-type SPOP localizes to liquid nuclear speckles, self-association-deficient SPOP mutants have a diffuse distribution in the nucleus. SPOP oligomerizes through its BTB and BACK domains. We show that BTB-mediated SPOP dimers form linear oligomers via BACK domain dimerization, and we determine the concentration-dependent populations of the resulting oligomeric species. Higher-order oligomerization of SPOP stimulates CRL3(SPOP) ubiquitination efficiency for its physiological substrate Gli3, suggesting that nuclear speckles are hotspots of ubiquitination. Dynamic, higher-order protein self-association may be a general mechanism to concentrate functional components in membrane-less cellular bodies.
authors.affiliation: Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN, USA.
authors.forename: Melissa R
authors.initials: MR
authors.lastname: Marzahn
authors.affiliation: Department of Cell and Molecular Biology, St. Jude Children's Research Hospital, Memphis, TN, USA.
authors.forename: Suresh
authors.initials: S
authors.lastname: Marada
authors.affiliation: Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN, USA.
authors.forename: Jihun
authors.initials: J
authors.lastname: Lee
authors.affiliation: Molecular Interaction Analysis Shared Resource, St. Jude Children's Research Hospital, Memphis, TN, USA.
authors.forename: Amanda
authors.initials: A
authors.lastname: Nourse
authors.affiliation: Wyatt Technology Corporation, Santa Barbara, CA, USA.
authors.forename: Sophia
authors.initials: S
authors.lastname: Kenrick
authors.affiliation: Dynamics of Macromolecular Assembly Section, Laboratory of Cellular Imaging and Macromolecular Biophysics, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Huaying
authors.initials: H
authors.lastname: Zhao
authors.affiliation: Department of Biological Chemistry, Weizmann Institute of Science, Rehovot, Israel.
authors.forename: Gili
authors.initials: G
authors.lastname: Ben-Nissan
authors.affiliation: Department of Cell and Molecular Biology, St. Jude Children's Research Hospital, Memphis, TN, USA.
authors.forename: Regina-Maria
authors.initials: RM
authors.lastname: Kolaitis
authors.affiliation: Department of Cellular Imaging Shared Resource, St. Jude Children's Research Hospital, Memphis, TN, USA.
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Peters
authors.affiliation: Department of Biostatistics, St. Jude Children's Research Hospital, Memphis, TN, USA.
authors.forename: Stanley
authors.initials: S
authors.lastname: Pounds
authors.affiliation: Department of Biochemistry, University of Toronto, Toronto, ON, Canada.
authors.forename: Wesley J
authors.initials: WJ
authors.lastname: Errington
authors.affiliation: Department of Biochemistry, University of Toronto, Toronto, ON, Canada Princess Margaret Cancer Centre, Toronto, ON, Canada.
authors.forename: Gilbert G
authors.initials: GG
authors.lastname: Privé
authors.affiliation: Department of Cell and Molecular Biology, Howard Hughes Medical Institute, St. Jude Children's Research Hospital, Memphis, TN, USA.
authors.forename: J Paul
authors.initials: JP
authors.lastname: Taylor
authors.affiliation: Department of Biological Chemistry, Weizmann Institute of Science, Rehovot, Israel.
authors.forename: Michal
authors.initials: M
authors.lastname: Sharon
authors.affiliation: Dynamics of Macromolecular Assembly Section, Laboratory of Cellular Imaging and Macromolecular Biophysics, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Peter
authors.initials: P
authors.lastname: Schuck
authors.affiliation: Department of Cell and Molecular Biology, St. Jude Children's Research Hospital, Memphis, TN, USA stacey.ogden@stjude.org tanja.mittag@stjude.org.
authors.forename: Stacey K
authors.initials: SK
authors.lastname: Ogden
authors.affiliation: Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN, USA stacey.ogden@stjude.org tanja.mittag@stjude.org.
authors.forename: Tanja
authors.identifier: 0000-0002-1827-3811
authors.initials: T
authors.lastname: Mittag
mesh_terms: D002467:Cell Nucleus / Q000378:metabolism*; D006801:Humans; D051741:Kruppel-Like Transcription Factors / Q000378:metabolism; D046911:Macromolecular Substances / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism*; D011485:Protein Binding; D000072417:Protein Domains; D055503:Protein Multimerization; D012097:Repressor Proteins / Q000378:metabolism*; D054875:Ubiquitination; D000074290:Zinc Finger Protein Gli3
chemical_list: C065683:GLI3 protein, human; D051741:Kruppel-Like Transcription Factors; D046911:Macromolecular Substances; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins; D012097:Repressor Proteins; C509761:SPOP protein, human; D000074290:Zinc Finger Protein Gli3
keywords: isodesmic self‐association; membrane‐less organelle; prostate cancer; speckle‐type POZ protein; ubiquitin ligase