title: Alternatively spliced insertions in the paired domain restrict the DNA sequence specificity of Pax6 and Pax8.
abstract: Transcription factors of the Pax family bind to their target genes via the paired domain which is known to be composed of two subdomains each recognizing distinct half-sites in adjacent major grooves of the DNA helix. We now demonstrate that the mammalian Pax8 gene gives rise, by alternative mRNA splicing, to a protein isoform containing an extra serine residue in the recognition alpha-helix 3 of the paired domain. This Pax8(S) protein does not interact with bipartite paired domain-binding sites, indicating that inactivation of the N-terminal DNA-binding motif severely restricts the sequence specificity of the paired domain. However, the Pax8(S) protein binds in vitro and in vivo to the 5aCON sequence which was previously identified as a high-affinity binding site for the Pax6(5a) splice variant carrying a 14-amino-acid insertion in the paired domain. The 5aCON sequence is shown to consist of four interdigitated 5' half-sites of the bipartite consensus sequence and is thus bound by four Pax8(S) molecules via the intact C-terminal DNA-binding motif of the paired domain. Together these data suggest that inactivation of the N-terminal region of the paired domain by alternative splicing is used in vivo to selectively target Pax transcription factors to gene regulatory regions containing highly specialized 5aCON-like sequences.
authors.affiliation: Research Institute of Molecular Pathology, Dr Bohr-Gasse 7, A-1030 Vienna, Austria.
authors.forename: Z
authors.initials: Z
authors.lastname: Kozmik
authors.forename: T
authors.initials: T
authors.lastname: Czerny
authors.forename: M
authors.initials: M
authors.lastname: Busslinger
mesh_terms: D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D016384:Consensus Sequence; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005136:Eye Proteins; D017930:Genes, Reporter; D018398:Homeodomain Proteins; D006801:Humans; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009687:Nuclear Proteins; D000071841:PAX6 Transcription Factor; D000072636:PAX8 Transcription Factor; D051761:Paired Box Transcription Factors; D011487:Protein Conformation; D012097:Repressor Proteins; D012694:Serine / Q000235:genetics; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: D004268:DNA-Binding Proteins; D005136:Eye Proteins; D018398:Homeodomain Proteins; D009687:Nuclear Proteins; D000071841:PAX6 Transcription Factor; C000605730:PAX6 protein, human; D000072636:PAX8 Transcription Factor; C083325:PAX8 protein, human; D051761:Paired Box Transcription Factors; C000605732:Pax6 protein, mouse; C499496:Pax8 protein, mouse; D012097:Repressor Proteins; D015534:Trans-Activators; D012694:Serine