DEVELOPMENTAL REGULATION AND STRUCTURAL ORGANIZATION OF CONNEXINS IN EPIDERMAL GAP-JUNCTIONS

The developmental regulation of gap junctions was analyzed in the deve loping rat epidermis by immunohistochemical and ultrastructural method s. The molecular composition of gap junction plaques was examined by l aser scanning confocal microscopy following immune-double labeling wit h monoclonal and polyclonal antibodies specific for alpha(1) (Cx43) an d beta(2) (Cx26) connexins, respectively. During early fetal developme nt (embryonic period), gap junctions were identified as large junction al plaques consisting of alpha(1) and beta(2) connexins. Ultrastructur ally, gap junctions were detected in the two-layered epidermis between the subapical borders of peridermal cells, at the periderm/basal laye r interface, and between the basal cells. The first ''switch'' in the utilization of alpha(1) and beta(2) connexins was observed at the onse t of epidermal stratification, when beta(2) expression was down-regula ted in the periderm and in the upper part of the intermedium. Gap junc tions were also detected ultrastructurally in all layers of the strati fied, nondifferentiated epidermis at E16. Junctional sizes included sm all plaques (0.05 mu m(2)) in the periderm, medium-size plaques (1 mu m(2)) in the upper part of the intermediate layer, and very large plaq ues (25 mu m(2)) in the basal layer. The second ''switch'' in the util ization of gap junction components coincided with epidermal differenti ation (>E18), when beta(2) was preferentially expressed in the differe ntiated granular and upper spinous layers. alpha(1) connexin was prese nt in the less differentiated spinous layer and in the proliferating b asal layer. Gap junctions were no longer detectable in the periderm fo llowing differentiation (keratinization) of the epidermis (E18-E20). A n analysis of immuno-double-stained sections by laser scanning confoca l microscopy revealed domains of potentially mixed and segregated anti gens within large junction plaques. These results indicated that large gap junction plaques (>1 pm in size) can contain segregated domains o f connexons, which contain a single protein (homooligomer). (C) 1994 A cademic Press, Inc.