title: Evaluation of the thyroid transcription factor-1 gene (TITF1) as a Hirschsprung's disease locus.
abstract: Hirschsprung's disease (HSCR, colonic aganglionosis) is an oligogenic entity that usually requires mutations in RET and other interacting loci. Decreased levels of RET expression may lead to the manifestation of HSCR. We previously showed that RET transcription was decreased due to alteration of the TITF1 binding site by two HSCR-associated RET promoter single nucleotide polymorphisms (SNPs). This prompted us to investigate whether DNA alterations in TITF1 could play a role in HSCR by affecting the RET-regulatory properties of the TITF1 protein. Our initial study on 86 Chinese HSCR patients revealed a Gly322Ser amino acid substitution in the TITF1protein. In this study we have examined an additional 102 Chinese and 70 Caucasian patients, and 194 Chinese and 60 Caucasian unselected, unrelated, subjects as controls. The relevance of the DNA changes detected in TITF1 by direct sequencing were evaluated using bioinformatics, reporter and binding-assays, mouse neurosphere culture, immunohistochemistry and immunofluorescence techniques. Met3Leu and Pro48Pro were identified in 2 Caucasian patients and 1 Chinese patient, respectively. In vitro analysis showed that Met3Leu reduced the activity of the RET promoter by 100% in the presence of the wild-type or HSCR-associated RET promoter SNP alleles. The apparent binding affinity of the TITF1 mutated protein was not decreased. The Met3Leu mutation may affect the interaction of TITF1 with its protein partners. The absence of Titf1 expression in mouse gut but not in human gut suggests that the role of TITF1 in gut development differs between the two species. TITF1 mutations could contribute to HSCR by affecting RET expression through defective interactions with other transcription factors.
authors.affiliation: Division of Paediatric Surgery, Department of Surgery, Li Ka Shing Faculty of Medicine of the University of Hong Kong, Hong Kong SAR, China.
authors.forename: Maria-Mercè
authors.initials: MM
authors.lastname: Garcia-Barceló
authors.forename: Danny Ko-chun
authors.initials: DK
authors.lastname: Lau
authors.forename: Elly Sau-wai
authors.initials: ES
authors.lastname: Ngan
authors.forename: Thomas Yuk-yu
authors.initials: TY
authors.lastname: Leon
authors.forename: Ting-ting
authors.initials: TT
authors.lastname: Liu
authors.forename: Man-ting
authors.initials: MT
authors.lastname: So
authors.forename: Xiao-ping
authors.initials: XP
authors.lastname: Miao
authors.forename: Vincent Chi-hang
authors.initials: VC
authors.lastname: Lui
authors.forename: Kenneth Kak-yuen
authors.initials: KK
authors.lastname: Wong
authors.forename: Raymond William
authors.initials: RW
authors.lastname: Ganster
authors.forename: Daniel Thomas
authors.initials: DT
authors.lastname: Cass
authors.forename: Geoffrey David Hain
authors.initials: GD
authors.lastname: Croaker
authors.forename: Paul Kwong-hang
authors.initials: PK
authors.lastname: Tam
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D044466:Asian People / Q000235:genetics; D001665:Binding Sites / Q000235:genetics; D016022:Case-Control Studies; D002460:Cell Line; D004247:DNA / Q000235:genetics / Q000378:metabolism; D005260:Female; D006367:HeLa Cells; D006627:Hirschsprung Disease / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008297:Male; D051379:Mice; D008808:Mice, Inbred CBA; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D017354:Point Mutation; D020641:Polymorphism, Single Nucleotide; D011401:Promoter Regions, Genetic; D017434:Protein Structure, Tertiary; D051096:Proto-Oncogene Proteins c-ret / Q000235:genetics; D000074482:Thyroid Nuclear Factor 1; D046509:Tissue Culture Techniques; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic; D044465:White People / Q000235:genetics
chemical_list: C000614412:NKX2-1 protein, human; C000614414:Nkx2-1 protein, mouse; D009687:Nuclear Proteins; D000074482:Thyroid Nuclear Factor 1; D014157:Transcription Factors; D004247:DNA; D051096:Proto-Oncogene Proteins c-ret; C099282:RET protein, human