We describe the creation of a transgenic zebrafish expressing GFP driven by a 7.5 kb promoter region of the tbx16 gene. This promoter segment is sufficient to recapitulate early embryonic expression of endogenous tbx16 in the presomitic mesoderm, the polster and, subsequently, in the hatching gland. Expression of GFP in the transgenic lines later in development diverges to some extent from endogenous tbx16 expression with the serendipitous result that one line expresses GFP specifically in commissural primary ascending (CoPA) interneurons of the developing spinal cord. Using this line we demonstrate that the gene mafba (valentino) is expressed in CoPA interneurons.
References
[1]
Chen JY, Chiou MJ (2010) Molecular cloning and functional analysis of the zebrafish luteinizing hormone beta subunit (LH) promoter. Fish Physiol Biochem 2010: 5.
[2]
Gong Z, Ju B, Wang X, He J, Wan H, et al. (2002) Green fluorescent protein expression in germ-line transmitted transgenic zebrafish under a stratified epithelial promoter from keratin8. Dev Dyn 223: 204–215.
[3]
Mione M, Baldessari D, Deflorian G, Nappo G, Santoriello C (2008) How neuronal migration contributes to the morphogenesis of the CNS: insights from the zebrafish. Dev Neurosci 30: 65–81.
[4]
Streisinger G, Walker C, Dower N, Knauber D, Singer F (1981) Production of clones of homozygous diploid zebra fish (Brachydanio rerio). Nature 291: 293–296.
[5]
Kimmel CB, Kane DA, Walker C, Warga RM, Rothman MB (1989) A mutation that changes cell movement and cell fate in the zebrafish embryo. Nature 337: 358–362.
[6]
Ho RK, Kane DA (1990) Cell-autonomous action of zebrafish spt-1 mutation in specific mesodermal precursors. Nature 348: 728–730.
[7]
Molven A, Wright CV, Bremiller R, De Robertis EM, Kimmel CB (1990) Expression of a homeobox gene product in normal and mutant zebrafish embryos: evolution of the tetrapod body plan. Development 109: 279–288.
[8]
Griffin KJ, Amacher SL, Kimmel CB, Kimelman D (1998) Molecular identification of spadetail: regulation of zebrafish trunk and tail mesoderm formation by T-box genes. Development 125: 3379–3388.
[9]
Ruvinsky I, Silver LM, Ho RK (1998) Characterization of the zebrafish tbx16 gene and evolution of the vertebrate T-box family. Dev Genes Evol 208: 94–99.
[10]
Tamme R, Wells S, Conran JG, Lardelli M (2002) The identity and distribution of neural cells expressing the mesodermal determinant spadetail. BMC Dev Biol 2: 9.
[11]
Wells S, Conran JG, Tamme R, Gaudin A, Webb J, et al. (2010) Cryptic organisation within an apparently irregular rostrocaudal distribution of interneurons in the embryonic zebrafish spinal cord. Exp Cell Res 2010: 1.
[12]
Westerfield M (2000) The zebrafish book. A guide for the laboratory use of zebrafish (Danio rerio). Eugene: University of Oregon Press.
[13]
Kimmel CB, Ballard WW, Kimmel SR, Ullmann B, Schilling TF (1995) Stages of embryonic development of the zebrafish. Dev Dyn 203: 253–310.
[14]
Jowett T (1997) Tissue In Situ Hybridization: Methods in Animal Development. New York: John Wiley & Sons, Inc. 128 p.
[15]
Ellingsen S, Laplante MA, Konig M, Kikuta H, Furmanek T, et al. (2005) Large-scale enhancer detection in the zebrafish genome. Development 132: 3799–3811.
[16]
Kuwada JY, Bernhardt RR, Nguyen N (1990) Development of spinal neurons and tracts in the zebrafish embryo. J Comp Neurol 302: 617–628.
[17]
Roberts A (2000) Early functional organization of spinal neurons in developing lower vertebrates. Brain Res Bull 53: 585–593.
[18]
Ju B, Chong SW, He J, Wang X, Xu Y, et al. (2003) Recapitulation of fast skeletal muscle development in zebrafish by transgenic expression of GFP under the mylz2 promoter. Dev Dyn 227: 14–26.
[19]
Park HC, Kim CH, Bae YK, Yeo SY, Kim SH, et al. (2000) Analysis of upstream elements in the HuC promoter leads to the establishment of transgenic zebrafish with fluorescent neurons. Dev Biol 227: 279–293.
[20]
Yeo SY, Kim M, Kim HS, Huh TL, Chitnis AB (2007) Fluorescent protein expression driven by her4 regulatory elements reveals the spatiotemporal pattern of Notch signaling in the nervous system of zebrafish embryos. Dev Biol 301: 555–567.
[21]
Kikuta H, Laplante M, Navratilova P, Komisarczuk AZ, Engstrom PG, et al. (2007) Genomic regulatory blocks encompass multiple neighboring genes and maintain conserved synteny in vertebrates. Genome Res 17: 545–555.
[22]
Higashijima S, Masino MA, Mandel G, Fetcho JR (2004) Engrailed-1 expression marks a primitive class of inhibitory spinal interneuron. J Neurosci 24: 5827–5839.
[23]
Kimura Y, Okamura Y, Higashijima S (2006) alx, a zebrafish homolog of Chx10, marks ipsilateral descending excitatory interneurons that participate in the regulation of spinal locomotor circuits. J Neurosci 26: 5684–5697.
