Entry created on 1 July 2019 (Revision 1.0) Annotator: Rita Pancsa; Bálint Mészáros; Orsolya Kovács
This entry is part of a multi-component system encompassing the following entries: O43791 Q9UER7
Basic protein information
Accession O43791
Common name SPOP
Gene SPOP
Organism Homo sapiens
Uniprot name Speckle-type POZ protein
Basic LLPS information
Organelle nuclear body; nuclear protein granule; SPOP/DAXX body
Type of experimental evidence
Joined entry O43791 Q9UER7
Protein region(s) mediating LLPS
28
-
359
Central region containing the ordered MATH (substrate binding) and BTB domains (dimerization and CUL3 binding)
Based on the experimental results of the following publication: 30244836
Molecular features viewer
PDB structures
View on PDBe
Extended LLPS information
Functional description
Mutations in the tumor suppressor SPOP (speckle-type POZ protein) cause prostate, breast and other solid tumors. SPOP is a substrate adaptor of the cullin3-RING ubiquitin ligase and localizes to nuclear speckles. Substrates trigger phase separation of SPOP in vitro and co-localization in membraneless organelles in cells. Substrates include the death-domain-associated protein (DAXX), androgen receptor (AR), and other important signaling cascade effectors, epigenetic modifiers and hormone signaling effectors, these contain multiple SPOP-binding (SB) motifs in their IDRs. Enzymatic activity correlates with cellular co-localization and in vitro mesoscale assembly formation. Disease-associated SPOP-mutations that lead to the accumulation of proto-oncogenic proteins interfere with phase separation and co-localization in membraneless organelles, suggesting that substrate-directed phase separation of this E3 ligase underlies the regulation of ubiquitin-dependent proteostasis (PMID:30244836).
Literature supporting the LLPS: 30244836
Functional class of membraneless organelle: activation/nucleation/signal amplification/bioreactor
Binding partners (at biological protein concentrations)
1) SPOP subtrates eg. DAXX or androgen receptor (strictly required for LLPS)
Type of RNA(s) required/used for the LLPS at biological protein concentrations
Not required.
Molecular interaction types contributing to LLPS
multivalent domain-motif interactions (PMID:30244836) linear oligomerization/self-association (PMID:27220849)
Determinants of phase separation and droplet properties
1) protein concentration of SPOP 2) substrate concentration 3) molar ratios SPOP:substrate 4) presence of cancer mutations (negatively affect LLPS)
Membrane cluster No
Partner-dependent Yes
RNA-dependent No
PTM required No
Domain-motif interactions Yes
Discrete oligomerization No
Regulation and disease
Post-translational modifications affecting LLPS
Position Residue PTM Effect Reference Modifying enzyme Notes
Isoforms known to affect LLPS
Isoform Effect Reference
All known isoforms containing sequence changes in the LLPS region(s)
Position type Isoform names from UniProt
Disease mutations affecting LLPS
Mutation dbSNP Disease OMIM Effect Reference Notes
Experimental information
Experimental techniques applied to prove/investigate LLPS
Transiently expressed mCherry-fused SPOP co-localized with GFP-fused DAXX (a SPOP substrate) in largely spherical type of nuclear bodies in vivo (morphology, protein localization) distinct from nuclear speckles, PML bodies, nucleoli, and Cajal bodies, as evidenced by flurescence microscopy. Different expression tags did not influence the co-localization of SPOP and DAXX. The co-expression of the two proteins change their localization, as SPOP localized to nuclear speckles, and DAXX localized to PML bodies when expressed alone. In vitro studies demonstrated that SPOP(28-359) undergoes self-oligomerization (physical interaction), and in the presence of molecular crowders such as Ficoll-70, these oligomers are large enough to be observed by light microscopy (particle size and count). At higher concentrations (change in protein concentration) in vitro DAXX(495-740) forms condensed droplets; however, this tendency is strongly enhanced in the presence of SPOP, and this was the case in the presence of both polymer and protein crowders, as evidenced by fluorescence microscopy of tagged protein constructs (PMID:30244836).
Experimental observations supporting the liquid material state of the condensate
morphological traits (PMID:30244836) rheological traits (PMID:30244836) dynamic movement/reorganization of molecules within the droplet assessed using FRAP (PMID:30244836)