CNBP is phosphorylated by Ser/Thr kinases during zebrafish embryonic development

LOMBARDO, V.; WEINER A.; ARMAS, P.; BORGOGNONE, M.; CALCATERRA, N. B.

Pinamar - Buenos Aires

Congreso; XLI Annual Meeting of the Argentine Society for Biochemistry and Molecular Biology (SAIB); X Congress of the Panamerican Association for Biochemistry and Molecular Biology (PABMB); XX Annual Meeting of the Argentine Society for Neurochemistry (SAN).; 2005

Sociedad Argentina de Investigación Bioquímica y Biología Molecular (SAIB); Panamerican Association for Biochemistry and Molecular Biology (PABMB); Sociedad Argentina de Neuroquímica (SAN)

Cellular nucleic acid binding protein (CNBP) is a small single-stranded nucleic acid binding protein with nucleic acid chaperone activity. It is made up of seven tandem CCHC-type zinc knuckle domains and an RGG box between the first and second zinc knuckles. CNBP primary structure shows a nuclear localization signal PKKEREQ and a PEST proteolytic site that yields two peptides of approximately 4000 and 15000 Da. Moreover, in silico analysis revealed the existence of several putative phosphorylation sites in Ser, Thr, and Tyr residues. We have previously shown that zebrafish CNBP is differentially phosphorylated both in vitro and in vivo during embryogensis. In this report, we show that a natural proteolytic CNBP product is differentially phosphorylated during early development, showing a phosphorylation pattern coincident with the one observed with the whole protein. We determined that CNBP is phosphorylated by Ser/Thr kinases present in zebrafish embryonic extracts. Using bidimentional electroforetic analysis, we observed that CNBP is phosphorylated in more than one amino acid residue, regardless the developmental stages used as source of kinase activity. To address which kinase is required to CNBP phosphorylation, we performed phosphorylation assays in presence of specific inhibitors of different kinases involved in defferentiation and cell proliferation.