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To link to full-text access for this article, visit this link: http://dx.doi.org/10.1016/j.ydbio.2007.11.034 Byline: Saradavey Vinothkumar, Sepand Rastegar, Masanari Takamiya, Raymond Ertzer, Uwe Strahle Keywords: Retina; Zebrafish; Enhancer; Shh; Gene regulation Abstract: The signaling molecule Sonic hedgehog (Shh) is required for differentiation of the vertebrate retina. In the developing zebrafish retina, shh expression is initiated at the ventronasal region, from where it spreads as a wave through the retina. To investigate the molecular mechanism underlying this coordinated expression of shh, we mapped the cis-regulatory region and identified a novel regulatory sequence in the first intron of the shh locus. This sequence contains binding sites for the transcription factors Erm and Pea3 that are known transducers of Fgf signaling. Mutation of the binding sites or knockdown of Pea3 and Erm abolishes transgene expression, indicating that Fgf signaling regulates shh expression in the retina. We provide evidence that Fgf3 and -8 control initiation of expression, while Fgf19 is crucial for the propagation of transgene expression through the retina. Inhibitor experiments indicate a continued requirement of FGF and Hedgehog (Hh) signaling for transgene expression after initiation at the ventronasal aspect of the retina. We propose a model, in which Fgf3 and -8 initiate expression and Fgf19 and Shh signals cooperate subsequently to promote establishment of expression throughout the retina. Author Affiliation: Institute for Toxicology and Genetics, Forschungszentrum Karlsruhe, Postfach 3640, 76021 Karlsruhe, Germany Article History: Received 7 September 2007; Revised 19 November 2007; Accepted 27 November 2007

To link to full-text access for this article, visit this link: http://dx.doi.org/10.1016/j.ydbio.2007.05.014 Byline: Christelle Etard (a)(b), Martine Behra (a)(d), Nadine Fischer (a), David Hutcheson (b), Robert Geisler (c), Uwe Strahle (a)(b) Keywords: Zebrafish; Chaperone; Co-chaperone; Muscle damage; hsp90a; UCS factor Abstract: Contraction of muscles is mediated by highly organized arrays of myosin motor proteins. We report here the characterization of a mutation of a UCS gene named steif/unc-45b that is required for the formation of ordered myofibrils in both the skeletal and cardiac muscles of zebrafish. We show that Steif/Unc-45b interacts with the chaperone Hsp90a in vitro. The two genes are co-expressed in the skeletal musculature and knockdown of Hsp90a leads to impaired myofibril formation in the same manner as lack of Steif/Unc-45b activity. Transcripts of both genes are up-regulated in steif mutants suggesting co-regulation of the two genes. Our data indicate a requirement of Steif/unc-45b and Hsp90a for the assembly of the contractile apparatus in the vertebrate skeletal musculature. Author Affiliation: (a) Institut de Genetique et de Biologie Moleculaire et Cellulaire, 1 rue Laurent Fries, 67404 Illkirch Cedex, CU de Strasbourg, France (b) Institute for Toxicology and Genetics, Forschungszentrum Karlsruhe, Postfach 3640, 76021 Karlsruhe, Germany (c) Max-Planck-Institut fur Entwicklungsbiologie, Spemannstr 35, D-72076 Tubingen, Germany (d) NIH/NHGRI Building 50, Bethesda, Maryland, USA Article History: Received 2 March 2007; Revised 9 May 2007; Accepted 15 May 2007

To link to full-text access for this article, visit this link: http://dx.doi.org/10.1016/j.ydbio.2006.11.004 Byline: Raymond Ertzer (a)(b), Ferenc Muller (a)(b), Yavor Hadzhiev (b), Saradavey Rathnam (b), Nadine Fischer (a), Sepand Rastegar (a)(b), Uwe Strahle (a)(b) Keywords: Embryonic shield; Notochord; Floor plate; Zona limitans intrathalamica; Hypothalamus; Zebrafish; Sonic hedgehog; Transgene; Enhancer Abstract: In zebrafish, as in other vertebrates, the secreted signalling molecule Sonic hedgehog (Shh) is expressed in organiser regions such as the embryonic midline and the zona limitans intrathalamica (zli). To investigate the regulatory mechanisms underlying the pattern of shh expression, we carried out a systematic analysis of the intronic regulatory sequences of zebrafish shh using stable transgenesis. Deletion analysis identified the modules responsible for expression in the embryonic shield, the hypothalamus and the zli and confirmed the activities of previously identified notochord and floor plate enhancers. We detected a strong synergism between regulatory regions. The degree of synergy varied over time in the hypothalamus suggesting different mechanisms for initiation and maintenance of expression. Our data show that the pattern of shh expression in the embryonic central nervous system involves an intricate crosstalk of at least 4 different regulatory regions. When compared to the enhancer activities of the mouse Shh gene, we observed a remarkable divergence of function of structurally conserved enhancer sequences. The activating region ar-C (61% identical to SFPE2 in mouse Shh), for example, mediates floor plate expression in the mouse embryo while it directs expression in the forebrain and the notochord and only weakly in the floor plate in the zebrafish embryo. This raises doubts on the predictive power of phylogenetic footprinting and indicates a stunning divergence of function of structurally conserved regulatory modules during vertebrate evolution. Author Affiliation: (a) Institut de Genetique et de Biologie Moleculaire et Cellulaire, CNRS/INSERM/ULP, BP 163, 67404 Illkirch Cedex, C.U. de Strasbourg, France (b) Institute of Toxicology and Genetics, Research Center Karlsruhe, PoB 3640, 76021 Karlsruhe, Germany Article History: Received 24 April 2006; Revised 30 October 2006; Accepted 4 November 2006

A study on the behavior of neural stem cells in the brain of zebrafish is discussed. Immunodetection of markers is done in the study.

The floor plate is an organising centre that controls neural differentiation and axonogenesis in the neural tube. The axon guidance molecule Netrin1 is expressed in the floor plate of zebrafish embryos. To elucidate the regulatory mechanisms underlying expression in the floor plate, we scanned the netrin1 locus for regulatory regions and identified an enhancer that drives expression in the floor plate and hypochord of transgenic embryos. The expression of the transgene is ectopically activated by Cyclops (Nodal) signals but does not respond to Hedgehog signals. The winged-helix transcription factor foxA2 (also HNF3[beta], axial) is expressed in the notochord and floor plate. We show that knock-down of FoxA2 leads to loss of floor plate, while notochord and hypochord development is unaffected, suggesting a specific requirement of FoxA2 in the floor plate. The transgene is ectopically activated by FoxA2, and expression of FoxA2 leads to rescue of floor plate differentiation in mutant embryos that are deficient in Cyclops signalling. Zebrafish and mouse use different signalling systems to specify floor plate. The zebrafish netrin1 regulatory region also drives expression in the floor plate of mouse and chicken embryos. This suggests that components of the regulatory circuits controlling expression in the floor plate are conserved and that FoxA2--given its importance for midline development also in the mouse--may be one such component. Key Words: cyclops; Nodal; floor plate; hypochord; netrin1; foxA2 (HNF3[beta], axial).