Identification and analysis of the zebrafish cnbp promoter and regulatory sequences.

WEINER, A; CALCATERRA, N.B.

Carlos Paz, Cordoba

Congreso; XLIV Reunion Annual – 44th Annual Meeting Argentine Society for Biochemistry and Molecular Biology.; 2008

SAIB

Cellular Nucleic acid Binding Protein (CNBP) is required for vertebrate rostral head development, by controlling the neural crest cells (NCC) proliferation/survival balance. Its embryonic gene expression is temporally and spatially restricted to NCC and some derivatives. To identify and elucidate zebrafish cnbp promoter as well as its regulatory mechanism, different genomic DNA fragments 5’ upstream the gene were cloned into the Tol2 transposon vector system. The characterization of cnbp minimal promoter was examined by following EGFP expression in live embryos and compared it with the reported embryonic gene expression pattern. As a result, two different regions were able to promote EGFP expression. One is located 800 bp upstream the putative start transcription site and displays a ubiquitous and low expression in the embryos. The other one, of 2100 bp, is surrounded by the intron 1 sequence and promotes a high and specific expression. Using different algorithms, we identified several putative promoter sequences and transcription factor binding sites (TFBS). Seven TFBS highly conserved among vertebrates were found. Five of them are not involved in embryonic development while the other two bind FOXD3 and PAX6, which are proteins responsible for normal NCC development. A hierarchical position of CNBP in the neural crest signalling pathway is proposed.