MID1 and MID2 homo- and heterodimerise to tether the rapamycin-sensitive PP2A regulatory subunit, alpha 4, to microtubules: implications for the clinical variability of X-linked Opitz GBBB syndrome and other developmental disorders.
BACKGROUND
Patients with Opitz GBBB syndrome present with a variable array of developmental defects including craniofacial, cardiac, and genital anomalies. Mutations in the X-linked MID1 gene, which encodes a microtubule-binding protein, have been found in approximately 50% of Opitz GBBB syndrome patients consistent with the genetically heterogeneous nature of the disorder. A protein highly related to MID1, called MID2, has also been described that similarly associates with microtubules.


RESULTS
To identify protein partners of MID1 and MID2 we undertook two separate yeast two-hybrid screens. Using this system we identified Alpha 4, a regulatory subunit of PP2-type phosphatases and a key component of the rapamycin-sensitive signaling pathway, as a strong interactor of both proteins. Analysis of domain-specific deletions has shown that the B-boxes of both MID1 and MID2 mediate the interaction with Alpha 4, the first demonstration in an RBCC protein of a specific role for the B-box region. In addition, we show that the MID1/2 coiled-coil motifs mediate both homo- and hetero-dimerisation, and that dimerisation is a prerequisite for association of the MID-Alpha 4 complex with microtubules.


CONCLUSIONS
Our findings not only implicate Alpha 4 in the pathogenesis of Opitz GBBB syndrome but also support our earlier hypothesis that MID2 is a modifier of the X-linked phenotype. Of further note is the observation that Alpha 4 maps to Xq13 within the region showing linkage to FG (Opitz-Kaveggia) syndrome. Overlap in the clinical features of FG and Opitz GBBB syndromes warrants investigation of Alpha 4 as a candidate for causing FG syndrome.
Short
Kieran M
KM
Department of Molecular Biosciences & ARC Special Research Centre for the Molecular Genetics of Development, University of Adelaide, Adelaide, South Australia, Australia 5005. kieran.short@adelaide.edu.au
Hopwood
Blair
B
Yi
Zou
Z
Cox
Timothy C
TC
D020816:Amino Acid Motifs / Q000235:genetics; D001665:Binding Sites; D019281:Dimerization; D040181:Genetic Diseases, X-Linked / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008025:Ligases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008868:Microtubule Proteins; D008869:Microtubule-Associated Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism*; D009687:Nuclear Proteins; D010749:Phosphoprotein Phosphatases / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D021122:Protein Subunits / Q000378:metabolism*; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D017384:Sequence Deletion / Q000235:genetics; D012694:Serine / Q000378:metabolism; D015398:Signal Transduction; D020123:Sirolimus / Q000378:metabolism*; D019082:Smith-Lemli-Opitz Syndrome / Q000235:genetics / Q000378:metabolism*; D013912:Threonine / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D044767:Ubiquitin-Protein Ligases
C121118:MID2 protein, human; D008868:Microtubule Proteins; D008869:Microtubule-Associated Proteins; D009687:Nuclear Proteins; D021122:Protein Subunits; D014157:Transcription Factors; D013912:Threonine; D012694:Serine; C109052:MID1 protein, human; D044767:Ubiquitin-Protein Ligases; D010749:Phosphoprotein Phosphatases; D008025:Ligases; D020123:Sirolimus