Functional redundancy of Tcf7 and Lef1, transcription factors mediating the Wnt signaling pathway, during zebrafish fin development

Gembu Abe, Saori Nagayoshi, Koichi Kawakami

National institute of Genetics, Sizuoka, Japan

   We have been developing the enhancer trap method in zebrafish by using the Tol2 transposable element. We created 73 fish lines that carried single copy insertions of the enhancer trap construct containing the zebrafish hsp70 promoter and the GFP gene and expressed GFP in specific cells, tissues and organs, and showed that the hsp70 promoter is highly capable of responding to chromosomal enhancers in zebrafish.
   One of these fish expressed GFP in the edge of the median fin fold and the pectoral fin during embryogenesis. In this line, the enhancer trap construct was integrated in the first exon of the tcf7 gene, which encodes a transcription factor mediating the Wnt canonical signaling pathway, and disrupted its function. Homozygous embryos showed shorter and wavy median and pectoral fins, indicating that Tcf7 plays a role in morphogenesis of these fins. It has been shown that Wnt signaling is essential for both limb/fin initiation and AER formation. However, the pectoral fins can still outgrow in the tcf7 homozygous embryos, and the mutant phenotype was weaker than that expected from a Wnt signaling deficiency in this region. In the pectoral fin bud, another member of the Tcf/Lef family, Lef1, is expressed. We knocked down the Lef1 activity by microinjection of lef1 MO into the tcf7 mutant. In the lef1 tcf7 double knock-down embryos, only rudimentary pectoral fins were formed and expression of the AER markers was abolished. Thus, Tcf7 and Lef1 are functionally redundant in AER formation during fin development.
   Even in the lef1 tcf7 double knock-down embryos, we found that fgf10 is induced in the mesenchyme, and outgrowth of the fin buds was still observed, indicating that fin initiation occurred. We hypothesized that another member of the Tcf/Lef family or a transcription factor that does not belong to this family is involved in mediating Wnt canonical signaling in this step, and currently testing this hypothesis.