Journal article
The Molecular Basis of Rieger Syndrome: ANALYSIS OF PITX2 HOMEODOMAIN PROTEIN ACTIVITIES
The Journal of biological chemistry, Vol.273(32), pp.20066-20072
08/07/1998
DOI: 10.1074/jbc.273.32.20066
PMID: 9685346
Abstract
Rieger syndrome is an autosomal-dominant developmental disorder that includes glaucoma and mild craniofacial dysmorphism in humans. Mutations in the Pitx2 homeobox gene have been linked to Rieger syndrome. We have characterized wild type and mutant Pitx2 activities using electrophoretic mobility shift assays, protein binding, and transient transfection assays. Pitx2 preferentially binds the bicoid homeodomain binding site and transactivates reporter genes containing this site. The combination of Pitx2 and another homeodomain protein, Pit-1, yielded a synergistic 55-fold activation of the prolactin promoter in transfection assays. Addition of Pit-1 increased Pitx2 binding to the bicoidelement in electrophoretic mobility shift assays. Furthermore, we demonstrate specific binding of Pit-1 to Pitx2 in vitro. Thus, wild type Pitx2 DNA binding activity is modulated by protein-protein interactions. We next studied two Rieger mutants. A threonine to proline mutation (T68P) in the second helix of the homeodomain retained DNA binding activity with the same apparentKD and only about a 2-fold reduction in theBmax. However, this mutant did not transactivate reporter genes containing the bicoid site. The mutant Pitx2 protein binds Pit-1, but there was no detectable synergism on the prolactin promoter. A second mutation (L54Q) in a highly conserved residue in helix 1 of the homeodomain yielded an unstable protein. Our results provide insights into the potential mechanisms underlying the developmental defects in Rieger syndrome.
Details
- Title: Subtitle
- The Molecular Basis of Rieger Syndrome: ANALYSIS OF PITX2 HOMEODOMAIN PROTEIN ACTIVITIES
- Creators
- Brad A AmendtLillian B SutherlandElena V SeminaAndrew F Russo
- Resource Type
- Journal article
- Publication Details
- The Journal of biological chemistry, Vol.273(32), pp.20066-20072
- DOI
- 10.1074/jbc.273.32.20066
- PMID
- 9685346
- NLM abbreviation
- J Biol Chem
- ISSN
- 0021-9258
- eISSN
- 1083-351X
- Language
- English
- Date published
- 08/07/1998
- Academic Unit
- Neurology; Orthodontics; Molecular Physiology and Biophysics; Anatomy and Cell Biology; Iowa Neuroscience Institute; Craniofacial Anomalies Research Center; Dental Research
- Record Identifier
- 9984020628502771
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