title: A noncoding point mutation of Zeb1 causes multiple developmental malformations and obesity in Twirler mice.
abstract: Heterozygous Twirler (Tw) mice develop obesity and circling behavior associated with malformations of the inner ear, whereas homozygous Tw mice have cleft palate and die shortly after birth. Zeb1 is a zinc finger protein that contributes to mesenchymal cell fate by repression of genes whose expression defines epithelial cell identity. This developmental pathway is disrupted in inner ears of Tw/Tw mice. The purpose of our study was to comprehensively characterize the Twirler phenotype and to identify the causative mutation. The Tw/+ inner ear phenotype includes irregularities of the semicircular canals, abnormal utricular otoconia, a shortened cochlear duct, and hearing loss, whereas Tw/Tw ears are severely malformed with barely recognizable anatomy. Tw/+ mice have obesity associated with insulin-resistance and have lymphoid organ hypoplasia. We identified a noncoding nucleotide substitution, c.58+181G>A, in the first intron of the Tw allele of Zeb1 (Zeb1(Tw)). A knockin mouse model of c.58+181G>A recapitulated the Tw phenotype, whereas a wild-type knockin control did not, confirming the mutation as pathogenic. c.58+181G>A does not affect splicing but disrupts a predicted site for Myb protein binding, which we confirmed in vitro. In comparison, homozygosity for a targeted deletion of exon 1 of mouse Zeb1, Zeb1(ΔEx1), is associated with a subtle abnormality of the lateral semicircular canal that is different than those in Tw mice. Expression analyses of E13.5 Twirler and Zeb1(ΔEx1) ears confirm that Zeb1(ΔEx1) is a null allele, whereas Zeb1(Tw) RNA is expressed at increased levels in comparison to wild-type Zeb1. We conclude that a noncoding point mutation of Zeb1 acts via a gain-of-function to disrupt regulation of Zeb1(Tw) expression, epithelial-mesenchymal cell fate or interactions, and structural development of the inner ear in Twirler mice. This is a novel mechanism underlying disorders of hearing or balance.
authors: Otolaryngology Branch, National Institute on Deafness and Other Communication Disorders, National Institutes of Health, Rockville, Maryland, United States of America.
authors: Kiyoto
authors: K
authors: Kurima
authors: Ronna
authors: R
authors: Hertzano
authors: Oksana
authors: O
authors: Gavrilova
authors: Kelly
authors: K
authors: Monahan
authors: Karl B
authors: KB
authors: Shpargel
authors: Garani
authors: G
authors: Nadaraja
authors: Yoshiyuki
authors: Y
authors: Kawashima
authors: Kyu Yup
authors: KY
authors: Lee
authors: Taku
authors: T
authors: Ito
authors: Yujiro
authors: Y
authors: Higashi
authors: David J
authors: DJ
authors: Eisenman
authors: Scott E
authors: SE
authors: Strome
authors: Andrew J
authors: AJ
authors: Griffith
mesh_terms: D000015:Abnormalities, Multiple / Q000235:genetics*; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D002352:Carrier Proteins / Q000235:genetics; D002874:Chromosome Mapping; D004268:DNA-Binding Proteins / Q000235:genetics; D007758:Ear, Inner / Q000002:abnormalities*; D058750:Epithelial-Mesenchymal Transition / Q000235:genetics; D018507:Gene Expression Regulation, Developmental; D055879:Gene Knock-In Techniques; D018398:Homeodomain Proteins / Q000235:genetics* / Q000378:metabolism*; D007438:Introns / Q000235:genetics*; D051741:Kruppel-Like Transcription Factors / Q000235:genetics* / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D009687:Nuclear Proteins / Q000235:genetics; D009765:Obesity / Q000235:genetics*; D010641:Phenotype; D017354:Point Mutation / Q000235:genetics; D022661:RNA, Untranslated / Q000235:genetics; D016601:RNA-Binding Proteins; D014157:Transcription Factors; D000071799:Zinc Finger E-box-Binding Homeobox 1
chemical_list: D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D051741:Kruppel-Like Transcription Factors; C110421:Mybbp1a protein, mouse; D009687:Nuclear Proteins; D022661:RNA, Untranslated; D016601:RNA-Binding Proteins; D014157:Transcription Factors; C519422:ZEB1 protein, mouse; D000071799:Zinc Finger E-box-Binding Homeobox 1