title: Identification of two independent SUMO-interacting motifs in Daxx: evolutionary conservation from Drosophila to humans and their biochemical functions.
abstract: Daxx is essential for embryonic development and implicated in apoptosis and transcriptional regulation. It is found only in the animal kingdom and appears to arise first in insects. In the Drosophila genus, the Daxx orthologs are much larger than those in other species. Here we show that in addition to a conserved core of approximately 200 residues, Daxx possesses several conserved domains and two essentially invariable short SUMO-interacting motifs (SIMs), each located at one or the other terminus of the protein. Both can independently interact with SUMO. The Daxx I7/733K double mutant with one mutation in each of the two SIMs no longer interacts with SUMO. Daxx interacts with Ubc9 and this interaction strictly requires at least one SIM. Interestingly, the Ubc9 H20D mutation that abolishes non-covalent Ubc9-SUMO interaction also interrupts Daxx-Ubc9 interaction. Thus, SUMO serves as the intermediate for Daxx-Ubc9 interaction. Surprisingly, Daxx I7/733K double mutant could still colocalize with PML. Furthermore, wt Daxx also strongly colocalizes with PMLDeltaS mutant, in which all three sumoylation sites are mutated, whereas PMLDeltaS only weakly colocalizes with Daxx I7/733K mutant, suggesting that SIM-SUMO interaction is not essential for but enhances PML-Daxx interaction. Remarkably, Daxx strongly stimulates c-Jun-mediated transcription and both SIMs are required for this stimulation. PML also activates c-Jun, which requires all three sumoylation sites. Coexpression of Daxx and PML revealed that they independently regulate c-Jun, with Daxx exerting a dominant role. These results suggest that the conserved SIMs are involved in mediating protein-protein interactions that underlie Daxx's diverse cellular functions.
authors.affiliation: Department of Anatomy and Cell Biology, Shands Cancer Center, University of Florida College of Medicine, 1376 Mowry Road, Gainesville, FL 32611, USA.
authors.forename: Aleixo
authors.initials: A
authors.lastname: Santiago
authors.forename: Adam C
authors.initials: AC
authors.lastname: Godsey
authors.forename: Jamil
authors.initials: J
authors.lastname: Hossain
authors.forename: Lisa Y
authors.initials: LY
authors.lastname: Zhao
authors.forename: Daiqing
authors.initials: D
authors.lastname: Liao
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry* / Q000378:metabolism*; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D056970:Co-Repressor Proteins; D017124:Conserved Sequence; D004331:Drosophila melanogaster / Q000378:metabolism*; D019143:Evolution, Molecular; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D018832:Molecular Chaperones; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000378:metabolism; D009687:Nuclear Proteins / Q000737:chemistry* / Q000378:metabolism*; D000071156:Promyelocytic Leukemia Protein; D011485:Protein Binding / Q000187:drug effects; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000187:drug effects; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D025841:Small Ubiquitin-Related Modifier Proteins / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D025521:Tumor Suppressor Proteins / Q000378:metabolism; D044763:Ubiquitin-Conjugating Enzymes / Q000378:metabolism; D000098843:Ubiquitin-Conjugating Enzyme UBC9
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D056970:Co-Repressor Proteins; C106985:DAXX protein, human; C526347:Daxx protein, Drosophila; D006877:Hydroxamic Acids; D018832:Molecular Chaperones; D050505:Mutant Proteins; D009687:Nuclear Proteins; D000071156:Promyelocytic Leukemia Protein; D016755:Proto-Oncogene Proteins c-jun; D025841:Small Ubiquitin-Related Modifier Proteins; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C070576:PML protein, human; C012589:trichostatin A; D044763:Ubiquitin-Conjugating Enzymes; D000098843:Ubiquitin-Conjugating Enzyme UBC9