Craniofacial Expression of PVRL1 in Early Embryogenesis in Zebrafish

Margarita Island Ectodermal Dysplasia is an autosomal recessive disorder affecting 1 in 2000 individuals on that island. The hallmark of this syndrome includes cleft lip, with or without cleft palate and ectodermal dysplasia(CLPED1). The gene responsible for this syndrome is believed to be loss of function of PVRL1 gene, which encodes nectin-1, an immunoglobulin related transmembrane cell-cell adhesion molecule (Suzuki et al. 2000). The zebrafish EST (AL925737) shows 68% and 75% identity with human and mouse PVRL1, respectively. Objective: The purpose of our study was to investigate zebrafish (Danio rerio) ortholog of human PVRL1 gene and its expression pattern in early embryogenesis in zebrafish. Method: RNA was extracted from one and two day embryos using TRIZOL Reagent (Invitrogen). SuperScript II RT-PCR (Invitrogen) was performed using specific primers for AL925737, yielding an approximate 500 base pair fragment. Expression of zebrafish AL925737 was detected by in situ hybridization, using a digoxigenin labeled riboprobe that spans the 500 base pair fragment. Results: Specific expression patterns were localized in the pharyngeal arches, forebrain, hindbrain and the retina of the eye. In two day embryos, the pharyngeal arches, the forebrain, the hindbrain and only one layer of the retina was expressed. However, in the three day embryos, there was another layer of expression in retina, however, pharyngeal arches expression was not noted. This pattern of the expression is similar to those of PVRL1 in Mus musculus. Conclusion: Our results indicate that PVRL1 homolog is present in zebrafish and expressed in Craniofacial region. Further investigations are in progress to delineate the exact pattern of the expression in pharyngeal arches region.