title: Aniridia-associated translocations, DNase hypersensitivity, sequence comparison and transgenic analysis redefine the functional domain of PAX6.
abstract: The transcription factor PAX6 plays a critical, evolutionarily conserved role in eye, brain and olfactory development. Homozygous loss of PAX6 function affects all expressing tissues and is neonatally lethal; heterozygous null mutations cause aniridia in humans and the Small eye (Sey) phenotype in mice. Several upstream and intragenic PAX6 control elements have been defined, generally through transgenesis. However, aniridia cases with chromosomal rearrangements far downstream of an intact PAX6 gene suggested a requirement for additional cis-acting control for correct gene expression. The likely location of such elements is pinpointed through YAC transgenic studies. A 420 kb yeast artificial chromosome (YAC) clone, extending well beyond the most distant patient breakpoint, was previously shown to rescue homozygous Small eye lethality and correct the heterozygous eye phenotype. We now show that a 310 kb YAC clone, terminating just 5' of the breakpoint, fails to influence the Sey phenotypes. Using evolutionary sequence comparison, DNaseI hypersensitivity analysis and transgenic reporter studies, we have identified a region, >150 kb distal to the major PAX6 promoter P1, containing regulatory elements. Components of this downstream regulatory region drive reporter expression in distinct partial PAX6 patterns, indicating that the functional PAX6 gene domain extends far beyond the transcription unit.
authors.affiliation: MRC Human Genetics Unit, Western General Hospital, Crewe Road, Edinburgh EH4 2XU, UK.
authors.forename: D A
authors.initials: DA
authors.lastname: Kleinjan
authors.forename: A
authors.initials: A
authors.lastname: Seawright
authors.forename: A
authors.initials: A
authors.lastname: Schedl
authors.forename: R A
authors.initials: RA
authors.lastname: Quinlan
authors.forename: S
authors.initials: S
authors.lastname: Danes
authors.forename: V
authors.initials: V
authors.lastname: van Heyningen
mesh_terms: D000818:Animals; D015783:Aniridia / Q000235:genetics*; D018244:Chromosomes, Artificial, Yeast / Q000235:genetics; D017931:DNA Primers / Q000737:chemistry; D003850:Deoxyribonuclease I / Q000378:metabolism*; D005136:Eye Proteins / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation / Q000235:genetics; D005809:Genes, Regulator / Q000502:physiology*; D017930:Genes, Reporter; D018398:Homeodomain Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D017403:In Situ Hybridization; D051379:Mice; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D008822:Mice, Transgenic; D000071841:PAX6 Transcription Factor; D051761:Paired Box Transcription Factors; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D012313:RNA / Q000378:metabolism; D012097:Repressor Proteins; D017422:Sequence Analysis, DNA; D014178:Translocation, Genetic
chemical_list: D017931:DNA Primers; D005136:Eye Proteins; D018398:Homeodomain Proteins; D000071841:PAX6 Transcription Factor; C000605730:PAX6 protein, human; D051761:Paired Box Transcription Factors; C000605732:Pax6 protein, mouse; D012097:Repressor Proteins; D012313:RNA; D003850:Deoxyribonuclease I