Involvement of Pax6 and Otx2 in the forebrain-specific regulation of the vertebrate homeobox gene ANF/Hesx1.
During early vertebrate development, ANF homeobox genes are expressed in the prospective forebrain. Their regulation is essential for correct morphogenesis and function of the prosencephalon. We identified a 1-kb fragment upstream of the chicken GANF gene sufficient to drive lacZ expression in the endogenous expression domain. Concordant with the high conservation of this sequence in five investigated species, this element is also active in the corresponding expression domain of the zebrafish orthologue. In vivo analysis of two in vitro-identified Otx2 binding sites in this conserved sequence revealed their necessity for activation of the chicken ANF promoter. In addition, we identified a Pax6-binding site close to the transcriptional start site that is occupied in vivo by Pax6 protein. Pax6 and GANF exhibit mutually exclusive expression domains in the anterior embryonic region. Overexpression of Pax6 in chick embryos inhibited the endogenous GANF expression, and in Pax6(-/-) mice the expression domain of the murine ANF orthologue Hesx1 was expanded and sustained, indicating inhibitory effects of Pax6 on GANF. However, a mutation of the Pax6 site did not abolish reporter activity from an electroporated vector. We conclude that Otx2 and Pax6 are key molecules involved in conserved mechanisms of ANF gene regulation.
Spieler
Derek
D
AG Entwicklungsbiologie, Max-Planck-Institut für biophysikalische Chemie, Am Fassberg, 37077 Göttingen, Germany.
Bäumer
Nicole
N
Stebler
Jürg
J
Köprunner
Marion
M
Reichman-Fried
Michal
M
Teichmann
Ulrike
U
Raz
Erez
E
Kessel
Michael
M
Wittler
Lars
L
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D051792:Basic Helix-Loop-Helix Proteins; D002642:Chick Embryo; D017124:Conserved Sequence; D005136:Eye Proteins; D018507:Gene Expression Regulation, Developmental; D005801:Genes, Homeobox; D018398:Homeodomain Proteins / Q000737:chemistry / Q000235:genetics* / Q000502:physiology*; D051379:Mice; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000502:physiology*; D051857:Otx Transcription Factors; D000071841:PAX6 Transcription Factor; D051761:Paired Box Transcription Factors; D011401:Promoter Regions, Genetic; D016548:Prosencephalon / Q000196:embryology* / Q000378:metabolism; D012097:Repressor Proteins; D015534:Trans-Activators / Q000502:physiology*; D000072056:Transcription Factor HES-1; D015027:Zebrafish; D029961:Zebrafish Proteins
D051792:Basic Helix-Loop-Helix Proteins; D005136:Eye Proteins; D018398:Homeodomain Proteins; D009419:Nerve Tissue Proteins; D051857:Otx Transcription Factors; D000071841:PAX6 Transcription Factor; D051761:Paired Box Transcription Factors; D012097:Repressor Proteins; D015534:Trans-Activators; D000072056:Transcription Factor HES-1; D029961:Zebrafish Proteins; C489736:HESX1 protein, human; C489734:Hes1 protein, mouse; C489733:Hesx1 protein, mouse; C497660:Otx2 protein, mouse; C000605730:PAX6 protein, human; C000605732:Pax6 protein, mouse; C497662:otx2b protein, zebrafish