title: MID1 catalyzes the ubiquitination of protein phosphatase 2A and mutations within its Bbox1 domain disrupt polyubiquitination of alpha4 but not of PP2Ac.
abstract: MID1 is a microtubule-associated protein that belongs to the TRIM family. MID1 functions as an ubiquitin E3 ligase, and recently was shown to catalyze the polyubiquitination of, alpha4, a protein regulator of protein phosphatase 2A (PP2A). It has been hypothesized that MID1 regulates PP2A, requiring the intermediary interaction with alpha4. Here we report that MID1 catalyzes the in vitro ubiquitination of the catalytic subunit of PP2A (PP2Ac) in the absence of alpha4. In the presence of alpha4, the level of PP2Ac ubiquitination is reduced. Using the MID1 RING-Bbox1-Bbox2 (RB1B2) construct containing the E3 ligase domains, we investigate the functional effects of mutations within the Bbox domains that are identified in patients with X-linked Opitz G syndrome (XLOS). The RB1B2 proteins harboring the C142S, C145T, A130V/T mutations within the Bbox1 domain and C195F mutation within the Bbox2 domain maintain auto-polyubiquitination activity. Qualitatively, the RB1B2 proteins containing these mutations are able to catalyze the ubiquitination of PP2Ac. In contrast, the RB1B2 proteins with mutations within the Bbox1 domain are unable to catalyze the polyubiquitination of alpha4. These results suggest that unregulated alpha4 may be the direct consequence of these natural mutations in the Bbox1 domain of MID1, and hence alpha4 could play a greater role to account for the increased amount of PP2A observed in XLOS-derived fibroblasts.
authors.affiliation: Department of Chemistry, George Washington University, Washington, District of Columbia, United States of America.
authors.forename: Haijuan
authors.initials: H
authors.lastname: Du
authors.affiliation: Department of Chemistry, George Washington University, Washington, District of Columbia, United States of America.
authors.forename: Kuanlin
authors.initials: K
authors.lastname: Wu
authors.affiliation: Department of Chemistry, George Washington University, Washington, District of Columbia, United States of America.
authors.forename: Alma
authors.initials: A
authors.lastname: Didoronkute
authors.affiliation: Department of Chemistry, George Washington University, Washington, District of Columbia, United States of America.
authors.forename: Marcus V A
authors.initials: MV
authors.lastname: Levy
authors.affiliation: Department of Chemistry, George Washington University, Washington, District of Columbia, United States of America.
authors.forename: Nimish
authors.initials: N
authors.lastname: Todi
authors.affiliation: Department of Chemistry, George Washington University, Washington, District of Columbia, United States of America.
authors.forename: Anna
authors.initials: A
authors.lastname: Shchelokova
authors.affiliation: Department of Chemistry, George Washington University, Washington, District of Columbia, United States of America.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Massiah
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D002972:Cleft Palate / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D004947:Esophagus / Q000002:abnormalities / Q000378:metabolism / Q000473:pathology; D005347:Fibroblasts / Q000378:metabolism / Q000473:pathology; D005786:Gene Expression Regulation; D040181:Genetic Diseases, X-Linked / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D006801:Humans; D006972:Hypertelorism / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D007021:Hypospadias / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008868:Microtubule Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008958:Models, Molecular; D018832:Molecular Chaperones; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D054648:Protein Phosphatase 2 / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D054875:Ubiquitination
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C490467:IGBP1 protein, human; D047908:Intracellular Signaling Peptides and Proteins; D008868:Microtubule Proteins; D018832:Molecular Chaperones; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C109052:MID1 protein, human; D044767:Ubiquitin-Protein Ligases; C517189:PPP2CA protein, human; D054648:Protein Phosphatase 2