title: Multiple enhancers located in a 1-Mb region upstream of POU3F4 promote expression during inner ear development and may be required for hearing.
abstract: POU3F4 encodes a POU-domain transcription factor required for inner ear development. Defects in POU3F4 function are associated with X-linked deafness type 3 (DFN3). Multiple deletions affecting up to ~900-kb upstream of POU3F4 are found in DFN3 patients, suggesting the presence of essential POU3F4 enhancers in this region. Recently, an inner ear enhancer was reported that is absent in most DFN3 patients with upstream deletions. However, two indications suggest that additional enhancers in the POU3F4 upstream region are required for POU3F4 function during inner ear development. First, there is at least one DFN3 deletion that does not eliminate the reported enhancer. Second, the expression pattern driven by this enhancer does not fully recapitulate Pou3f4 expression in the inner ear. Here, we screened a 1-Mb region upstream of the POU3F4 gene for additional cis-regulatory elements and searched for novel DFN3 mutations in the identified POU3F4 enhancers. We found several novel enhancers for otic vesicle expression. Some of these also drive expression in kidney, pancreas and brain, tissues that are known to express Pou3f4. In addition, we report a new and smallest deletion identified so far in a DFN3 family which eliminates 3.9 kb, comprising almost exclusively the previous reported inner ear enhancer. We suggest that multiple enhancers control the expression of Pou3f4 in the inner ear and these may contribute to the phenotype observed in DFN3 patients. In addition, the novel deletion demonstrates that the previous reported enhancer, although not sufficient, is essential for POU3F4 function during inner ear development.
authors.affiliation: Centro Andaluz de Biología del Desarrollo, Consejo Superior de Investigaciones Científicas, Universidad Pablo de Olavide, Carretera de Utrera Km1, 41013, Sevilla, Spain.
authors.forename: Silvia
authors.initials: S
authors.lastname: Naranjo
authors.forename: Krysta
authors.initials: K
authors.lastname: Voesenek
authors.forename: Elisa
authors.initials: E
authors.lastname: de la Calle-Mustienes
authors.forename: Alex
authors.initials: A
authors.lastname: Robert-Moreno
authors.forename: Haris
authors.initials: H
authors.lastname: Kokotas
authors.forename: Maria
authors.initials: M
authors.lastname: Grigoriadou
authors.forename: John
authors.initials: J
authors.lastname: Economides
authors.forename: Guy
authors.initials: G
authors.lastname: Van Camp
authors.forename: Nele
authors.initials: N
authors.lastname: Hilgert
authors.forename: Felipe
authors.initials: F
authors.lastname: Moreno
authors.forename: Berta
authors.initials: B
authors.lastname: Alsina
authors.forename: Michael B
authors.initials: MB
authors.lastname: Petersen
authors.forename: Hannie
authors.initials: H
authors.lastname: Kremer
authors.forename: José Luis
authors.initials: JL
authors.lastname: Gómez-Skarmeta
mesh_terms: D024506:5' Flanking Region / Q000235:genetics*; D000818:Animals; D001483:Base Sequence; D004252:DNA Mutational Analysis; D007758:Ear, Inner / Q000254:growth & development / Q000378:metabolism*; D004625:Embryo, Nonmammalian / Q000196:embryology / Q000378:metabolism; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D005192:Family Health; D005260:Female; D017353:Gene Deletion; D018507:Gene Expression Regulation, Developmental; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006309:Hearing / Q000235:genetics*; D034381:Hearing Loss / Q000235:genetics; D006801:Humans; D017403:In Situ Hybridization; D008297:Male; D008856:Microscopy, Fluorescence; D050809:POU Domain Factors / Q000235:genetics*; D010375:Pedigree; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D014981:Xenopus / Q000196:embryology / Q000235:genetics
chemical_list: D050809:POU Domain Factors; C091774:POU3F4 protein, human; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins