The tight junctions of the endothelial and epithelial regions of the blood-brain barrier are regulated by interactions with the neighboring tissue. We examined how the neural retina regulates the assembly of tight junctions in the retinal pigment epithelium (RPE). The proteins JAM-A, AF-6, PAR-3 and PAR-6 have been implicated in the assembly of other epithelial tight junctions. Using chick embryos and primary cell culture, we examined gene expression of these proteins during embryonic development, and whether retinal secretions regulate their expression. Three highly conserved RNA splice sites of AF-6 were identified in chick ocular tissues, but only two were expressed in RPE. JAM-A and AF-6 were expressed at relatively high levels early in development when adherens junctions form, but before tight junctions form. Expression of JAM-A and the AF-6 isoforms actually decreased when tight junctions were forming and expanding. The expression of PAR-3 and PAR-6 was constant. Despite the expression of these proteins in vitro (along with claudins, occludin, ZO-1 and ZO-2), the tight junctional networks that form were discontinuous (Rahner, C., Fukuhara, M., Peng, S., Kojima, S., Rizzolo, L.J., 2004. The apical and basal environments of the retinal pigment epithelium regulate the maturation of tight junctions during development. J. Cell Sci. 117, 3307-3318). The expression of these assembly proteins was unaffected by a retinal conditioned medium that induced the completion of tight junction formation. These data indicate that the early expression of the assembly proteins corresponds to the initial establishment of the adherens and tight junctions, but secretory products of the neural retina must induce the expression of additional proteins to complete the maturation process.