title: Hoxb3 negatively regulates Hoxb1 expression in mouse hindbrain patterning.
abstract: The spatial regulation of combinatorial expression of Hox genes is critical for determining hindbrain rhombomere (r) identities. To address the cross-regulatory relationship between Hox genes in hindbrain neuronal specification, we have generated a gain-of-function transgenic mouse mutant Hoxb3(Tg) using the Hoxb2 r4-specific enhancer element. Interestingly, in r4 of the Hoxb3(Tg) mutant where Hoxb3 was ectopically expressed, the expression of Hoxb1 was specifically abolished. The hindbrain neuronal defects of the Hoxb3(Tg) mutant mice were similar to those of Hoxb1(-/-) mutants. Therefore, we hypothesized that Hoxb3 could directly suppress Hoxb1 expression. We first identified a novel Hoxb3 binding site S3 on the Hoxb1 locus and confirmed protein binding to this site by EMSA, and by in vivo ChIP analysis using P19 cells and hindbrain tissues from the Hoxb3(Tg) mutant. We further showed that Hoxb3 could suppress Hoxb1 transcriptional activity by chick in ovo luciferase reporter assay. Moreover, in E10.5 wildtype caudal hindbrain, where Hoxb1 is not expressed, we showed by in vivo ChIP that Hoxb3 was consistently bound to the S3 site on the Hoxb1 gene. This study reveals a novel negative regulatory mechanism by which Hoxb3 as a posterior gene serves to restrict Hoxb1 expression in r4 by direct transcriptional repression to maintain the rhombomere identity.
authors.affiliation: Department of Biochemistry, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong, China.
authors.forename: Elaine Y M
authors.initials: EY
authors.lastname: Wong
authors.forename: Xing An
authors.initials: XA
authors.lastname: Wang
authors.forename: Siu Shan
authors.initials: SS
authors.lastname: Mak
authors.forename: Jearn Jang
authors.initials: JJ
authors.lastname: Sae-Pang
authors.forename: Kam Wing
authors.initials: KW
authors.lastname: Ling
authors.forename: Bernd
authors.initials: B
authors.lastname: Fritzsch
authors.forename: Mai Har
authors.initials: MH
authors.lastname: Sham
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D030161:Avian Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D019521:Body Patterning; D002642:Chick Embryo; D019465:Craniofacial Abnormalities / Q000196:embryology / Q000235:genetics / Q000378:metabolism; D017931:DNA Primers / Q000235:genetics; D018507:Gene Expression Regulation, Developmental; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008817:Mice, Mutant Strains; D008822:Mice, Transgenic; D008959:Models, Neurological; D055495:Neurogenesis / Q000235:genetics / Q000502:physiology; D012249:Rhombencephalon / Q000196:embryology* / Q000378:metabolism*
chemical_list: D030161:Avian Proteins; D017931:DNA Primers; C101121:HOXB1 homeodomain protein; D018398:Homeodomain Proteins; C503713:Hoxb3 protein, mouse