Identification of Pax2-regulated genes by expression profiling of the mid-hindbrain organizer region.
The paired domain transcription factor Pax2 is required for the formation of the isthmic organizer (IsO) at the midbrain-hindbrain boundary, where it initiates expression of the IsO signal Fgf8. To gain further insight into the role of Pax2 in mid-hindbrain patterning, we searched for novel Pax2-regulated genes by cDNA microarray analysis of FACS-sorted GFP+ mid-hindbrain cells from wild-type and Pax2-/- embryos carrying a Pax2(GFP) BAC transgene. Here, we report the identification of five genes that depend on Pax2 function for their expression in the mid-hindbrain boundary region. These genes code for the transcription factors En2 and Brn1 (Pou3f3), the intracellular signaling modifiers Sef and Tapp1, and the non-coding RNA Ncrms. The Brn1 gene was further identified as a direct target of Pax2, as two functional Pax2-binding sites in the promoter and in an upstream regulatory element of Brn1 were essential for lacZ transgene expression at the mid-hindbrain boundary. Moreover, ectopic expression of a dominant-negative Brn1 protein in chick embryos implicated Brn1 in Fgf8 gene regulation. Together, these data defined novel functions of Pax2 in the establishment of distinct transcriptional programs and in the control of intracellular signaling during mid-hindbrain development.
Bouchard
Maxime
M
Research Institute of Molecular Pathology, Vienna Biocenter, Dr Bohr-Gasse 7, 1030 Vienna, Austria. maxime.bouchard@mcgill.ca
Grote
David
D
Craven
Sarah E
SE
Sun
Qiong
Q
Steinlein
Peter
P
Busslinger
Meinrad
M
D000818:Animals; D002642:Chick Embryo; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D018274:Electroporation; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D049452:Green Fluorescent Proteins; D018398:Homeodomain Proteins / Q000378:metabolism; D017403:In Situ Hybridization; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D007763:Lac Operon / Q000235:genetics; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice / Q000196:embryology*; D008822:Mice, Transgenic; D009419:Nerve Tissue Proteins / Q000378:metabolism; D009479:Neuropeptides / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D020897:Organizers, Embryonic / Q000196:embryology* / Q000378:metabolism; D051762:PAX2 Transcription Factor; D050809:POU Domain Factors; D012249:Rhombencephalon / Q000196:embryology* / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D015534:Trans-Activators / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D019076:Transgenes / Q000235:genetics; D000099266:Engrailed 2 Protein
D017931:DNA Primers; D004268:DNA-Binding Proteins; D049452:Green Fluorescent Proteins; D018398:Homeodomain Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D009479:Neuropeptides; D051762:PAX2 Transcription Factor; D050809:POU Domain Factors; D015534:Trans-Activators; D014157:Transcription Factors; D000099266:Engrailed 2 Protein; C497319:Pax2 protein, mouse; C461495:Plekha1 protein, mouse; C469338:Sef protein, mouse; C074065:Pou3f3 protein, mouse