The Signalling Molecule Sonic Hedgehog Is Involved in a Multitude of Distinct Patterning Processes during Vertebrate Embryogenesis. In the Nascent Body Axis of the Zebrafish Embryo, Sonic Hedgehog Is Co-Expressed with Axial (Hnf3b in Mammals), a Transcrip
Resource
DEVELOPMENT v.126 n.10 pp.2103-2116
Journal
DEVELOPMENT
Journal Volume
v.126
Journal Issue
n.10
Pages
2103-2116
Date Issued
1999
Date
1999
Author(s)
MULLER, F.
CHANG, BEI-EN
Abstract
The signalling molecule Sonic hedgehog (Shh) controls a wide range of differentiation processes during vertebrate development. Numerous studies have suggested that the absolute levels as well as correct spatial and temporal expression of shh are critical for its function. To investigate the regulation of shh expression, we have studied the mechanism controlling its spatial expression in the zebrafish. We employed an enhancer screening strategy in zebrafish embryos based on co-injection of putative enhancer sequences with a reporter construct and analysis of mosaic expression in accumulated expression maps. Enhancers were identified in intron 1 and 2 that mediate floor plate and notochord expression. These enhancers also drive notochord and floor plate expression in the mouse embryo strongly suggesting that the mechanisms controlling shh expression in the midline are conserved between zebrafish and mouse. Functional analysis in the zebrafish embryo revealed that the intronic enhancers have a complex organisation. Two activator regions, ar- A and ar-C, were identified in intron 1 and 2, respectively, which mediate mostly notochord and floor plate expression. In contrast, another activating region, ar-B, in intron 1 drives expression in the floor plate. Deletion fine mapping of ar-C delineated three regions of 40 bp to be essential for activity. These regions do not contain binding sites for HNF 3b, the winged helix transcription factor previously implicated in the regulation of shh expression, indicating the presence of novel regulatory mechanisms. A T-box transcription factor-binding site was found in a functionally important region that forms specific complexes with protein extracts from wild-type but not from notochord- deficient mutant embryos.
Subjects
floor plate, notochord
sonic hedgehog
HNF3b
enhancer
gene regulation
zebrafish
mouse transgenic
co-injection
T-box