title: Requirement of PML SUMO interacting motif for RNF4- or arsenic trioxide-induced degradation of nuclear PML isoforms.
abstract: PML, the organizer of nuclear bodies (NBs), is expressed in several isoforms designated PMLI to VII which differ in their C-terminal region due to alternative splicing of a single gene. This variability is important for the function of the different PML isoforms. PML NB formation requires the covalent linkage of SUMO to PML. Arsenic trioxide (As₂O₃) enhances PML SUMOylation leading to an increase in PML NB size and promotes its interaction with RNF4, a poly-SUMO-dependent ubiquitin E3 ligase responsible for proteasome-mediated PML degradation. Furthermore, the presence of a bona fide SUMO Interacting Motif (SIM) within the C-terminal region of PML seems to be required for recruitment of other SUMOylated proteins within PML NBs. This motif is present in all PML isoforms, except in the nuclear PMLVI and in the cytoplasmic PMLVII. Using a bioluminescence resonance energy transfer (BRET) assay in living cells, we found that As₂O₃ enhanced the SUMOylation and interaction with RNF4 of nuclear PML isoforms (I to VI). In addition, among the nuclear PML isoforms, only the one lacking the SIM sequence, PMLVI, was resistant to As₂O₃-induced PML degradation. Similarly, mutation of the SIM in PMLIII abrogated its sensitivity to As₂O₃-induced degradation. PMLVI and PMLIII-SIM mutant still interacted with RNF4. However, their resistance to the degradation process was due to their inability to be polyubiquitinated and to recruit efficiently the 20S core and the β regulatory subunit of the 11S complex of the proteasome in PML NBs. Such resistance of PMLVI to As₂O₃-induced degradation was alleviated by overexpression of RNF4. Our results demonstrate that the SIM of PML is dispensable for PML SUMOylation and interaction with RNF4 but is required for efficient PML ubiquitination, recruitment of proteasome components within NBs and proteasome-dependent degradation of PML in response to As₂O₃.
authors.affiliation: CNRS, FRE 3235, Université Paris Descartes, Paris, France.
authors.forename: Mohamed Ali
authors.initials: MA
authors.lastname: Maroui
authors.forename: Sabrina
authors.initials: S
authors.lastname: Kheddache-Atmane
authors.forename: Faten
authors.initials: F
authors.lastname: El Asmi
authors.forename: Laurent
authors.initials: L
authors.lastname: Dianoux
authors.forename: Muriel
authors.initials: M
authors.lastname: Aubry
authors.forename: Mounira K
authors.initials: MK
authors.lastname: Chelbi-Alix
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D000077237:Arsenic Trioxide; D001152:Arsenicals / Q000494:pharmacology*; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism*; D003593:Cytoplasm / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D051379:Mice; D009154:Mutation; D009687:Nuclear Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D010087:Oxides / Q000494:pharmacology*; D000071156:Promyelocytic Leukemia Protein; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D059748:Proteolysis / Q000187:drug effects*; D025842:SUMO-1 Protein / Q000378:metabolism*; D058207:Sumoylation / Q000187:drug effects; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D025521:Tumor Suppressor Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D001152:Arsenicals; D009687:Nuclear Proteins; D010087:Oxides; C513293:Pml protein, mouse; D000071156:Promyelocytic Leukemia Protein; D020033:Protein Isoforms; C114070:RNF4 protein, human; D025842:SUMO-1 Protein; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D046988:Proteasome Endopeptidase Complex; D000077237:Arsenic Trioxide