The Opitz syndrome gene product MID1 assembles a microtubule-associated ribonucleoprotein complex.
Opitz BBB/G syndrome (OS) is a heterogenous malformation syndrome mainly characterised by hypertelorism and hypospadias. In addition, patients may present with several other defects of the ventral midline such as cleft lip and palate and congenital heart defects. The syndrome-causing gene encodes the X-linked E3 ubiquitin ligase MID1 that mediates ubiquitin-specific modification and degradation of the catalytic subunit of the translation regulator protein phosphatase 2A (PP2A). Here, we show that the MID1 protein also associates with elongation factor 1alpha (EF-1alpha) and several other proteins involved in mRNA transport and translation, including RACK1, Annexin A2, Nucleophosmin and proteins of the small ribosomal subunits. Mutant MID1 proteins as found in OS patients lose the ability to interact with EF-1alpha. The composition of the MID1 protein complex was determined by several independent methods: (1) yeast two-hybrid screening and (2) immunofluorescence, (3) a biochemical approach involving affinity purification of the complex, (4) co-fractionation in a microtubule assembly assay and (5) immunoprecipitation. Moreover, we show that the cytoskeleton-bound MID1/translation factor complex specifically associates with G- and U-rich RNAs and incorporates MID1 mRNA, thus forming a microtubule-associated ribonucleoprotein (RNP) complex. Our data suggest a novel function of the OS gene product in directing translational control to the cytoskeleton. The dysfunction of this mechanism would lead to malfunction of microtubule-associated protein translation and to the development of OS.
Aranda-Orgillés
Beatriz
B
Max-Planck Institute for Molecular Genetics, Ihnestr. 73, 14195, Berlin, Germany. aranda@molgen.mpg.de
Trockenbacher
Alexander
A
Winter
Jennifer
J
Aigner
Johanna
J
Köhler
Andrea
A
Jastrzebska
Ewa
E
Stahl
Joachim
J
Müller
Eva-Christina
EC
Otto
Albrecht
A
Wanker
Erich E
EE
Schneider
Rainer
R
Schweiger
Susann
S
D017306:Annexin A2 / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D002846:Chromatography, Affinity; D005455:Fluorescent Antibody Technique; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism; D040181:Genetic Diseases, X-Linked / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D017403:In Situ Hybridization; D008868:Microtubule Proteins / Q000235:genetics* / Q000378:metabolism; D008870:Microtubules / Q000378:metabolism*; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D000090243:Nucleophosmin; D020648:Peptide Elongation Factor 1 / Q000235:genetics / Q000378:metabolism*; D012313:RNA / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000494:pharmacology; D000075370:Receptors for Activated C Kinase; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism; D012261:Ribonucleoproteins / Q000235:genetics / Q000378:metabolism*; D012689:Sequence Homology, Nucleic Acid; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D020798:Two-Hybrid System Techniques; D044767:Ubiquitin-Protein Ligases
C472822:ANXA2 protein, human; D017306:Annexin A2; D008868:Microtubule Proteins; C000716967:NPM1 protein, human; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D020648:Peptide Elongation Factor 1; C419368:RACK1 protein, human; D034741:RNA, Small Interfering; D000075370:Receptors for Activated C Kinase; D011956:Receptors, Cell Surface; D012261:Ribonucleoproteins; D014157:Transcription Factors; D000090243:Nucleophosmin; D012313:RNA; C109052:MID1 protein, human; D044767:Ubiquitin-Protein Ligases; D019204:GTP-Binding Proteins