Pax-2 regulatory sequences that direct transgene expression in the developing neural plate and external granule cell layer of the cerebellum.
Expression of Pax-2 in the mouse gastrula is the first marker of the midbrain-hindbrain region. To address roles played by transcription factors in the process of neural plate pattern formation and to facilitate gain-of-function approaches in the study of midbrain-hindbrain and cerebellar development, we characterized regulatory sequences at the Pax-2 locus using an in vivo transgenic mouse reporter assay. An 8.5 kb fragment of genomic DNA located upstream of Pax-2 directed lacZ expression prior to neurulation (7.5 days post-coitum, dpc) in a region fated to become midbrain and hindbrain, and subsequently in developing neuroepithelium. While similar to the pattern of Pax-2 expression, reporter gene activity extended beyond the boundaries of Pax-2 expression, most probably reflecting purdurance of beta-galactosidase activity and an absence of DNA sequences that restrict Pax-2 expression to rhombomere 1 by 9. 5 dpc. In the fetal and neonatal brain, Pax-2-lacZ activity was confined largely to Purkinje cells and the external granule cell layer (EGL) of the cerebellum. A 4 kb regulatory element, in contrast, initiated neural expression at 8.25 dpc in the anterior hindbrain, but recapitulated all later aspects of Pax-2-lacZ activity observed with the larger transgene. These results indicate the presence of regulatory sequences upstream of the Pax-2 locus capable of directing gene expression in the developing midbrain, first rhombomere of the hindbrain, and its principal derivative, the cerebellum. Successful misexpression of Sonic hedgehog demonstrates that Pax-2 regulatory sequences should prove generally useful for transgenic gain-of-function approaches in mice.
Rowitch
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DH
Department of Molecular and Cellular Biology, Harvard University, 16 Divinity Avenue, Cambridge, MA 02138, USA. david_rowitch@dfci.harvard.edu
Kispert
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A
McMahon
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AP
D000818:Animals; D019521:Body Patterning / Q000235:genetics; D002531:Cerebellum / Q000166:cytology / Q000196:embryology* / Q000378:metabolism*; D018076:DNA, Complementary / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics*; D005260:Female; D005775:Gastrula / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D017930:Genes, Reporter / Q000235:genetics; D053823:Hedgehog Proteins; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D008822:Mice, Transgenic; D051762:PAX2 Transcription Factor; D011506:Proteins / Q000235:genetics; D020218:Response Elements / Q000235:genetics*; D019170:Somites / Q000378:metabolism; D013997:Time Factors; D015534:Trans-Activators; D014157:Transcription Factors / Q000235:genetics*; D015533:Transcriptional Activation / Q000235:genetics; D019076:Transgenes / Q000235:genetics*
D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D053823:Hedgehog Proteins; D051762:PAX2 Transcription Factor; C497319:Pax2 protein, mouse; D011506:Proteins; D015534:Trans-Activators; D014157:Transcription Factors