Abstract•

No abstract

The signal transduction adapter protein Disabled-2 (Dab2) is one of the two mammalian orthologs of the Drosophila Disabled. The brain-specific Disabled-1 (Dab1) functions in positional organization of brain cells during development. Dab2 is widely distributed and is highly expressed in many epithelial cell types. The dab2 gene was interrupted by in-frame insertion of beta-galactosidase (LacZ) in embryonic stem cells and transgenic mice were produced. Dab2 expression was first observed in the primitive endoderm at E4.5, immediately following implantation. The homozygous Dab2-deficient mutant is embryonic lethal (earlier than E6.5) due to defective cell positioning and structure formation of the visceral endoderm. In E5.5 dab2 (-/-) conceptus, visceral endoderm-like cells are present in the deformed primitive egg cylinder; however, the visceral endoderm cells are not organized, the cells of the epiblast have not expanded, and the proamniotic cavity fails to form. Disorganization of the visceral endodermal layer is evident, as cells with positive visceral endoderm markers are scattered throughout the dab2 (-/-) conceptus. Only degenerated remains were observed at E6.5 for dab2 (-/-) embryos, and by E7.5, the defective embryos were completely reabsorbed. In blastocyst in vitro culture, initially cells with characteristics of endoderm, trophectoderm, and inner cell mass were observed in the outgrowth of the hatched dab2 (-/-) blastocysts. However, the dab2 (-/-) endodermal cells are much more dispersed and disorganized than those from wild-type blastocysts, the inner cell mass fails to expand, and the outgrowth degenerates by day 7. Thus, Dab2 is required for visceral endodermal cell organization during early mouse development. The absence of an organized visceral endoderm in Dab2-deficient conceptus leads to the growth failure of the inner cell mass. We suggest that Dab2 functions in a signal pathway to regulate endodermal cell organization using endocytosis of ligands from the blastocoel cavity as a positioning cue.

The formation of the primitive endoderm layer on the surface of the inner cell mass is one of the earliest epithelial morphogenesis in mammalian embryos. In mouse embryos deficient of Disabled-2 (Dab2), the primitive endoderm cells lose the ability to position on the surface, resulting in defective morphogenesis. Embryonic stem cells lacking Dab2 are also unable to position on the surface of cell aggregates and fail to form a primitive endoderm outer layer in the embryoid bodies. The cellular function of Dab2, a cargo-selective adaptor, in mediating endocytic trafficking of clathrin-coated vesicles is well established. We show here that Dab2 mediates directional trafficking and polarized distribution of cell surface proteins such as megalin and E-cadherin and propose that loss of polarity is the underlying mechanism for the loss of epithelial cell surface positioning in Dab2-deficient embryos and embryoid bodies. Thus, the findings indicate that Dab2 is a surface positioning gene and suggest a novel mechanism of epithelial cell surface targeting.Epithelial cells are positioned on the outer surface of organs or the inner surface of glandular structures and are involved in diverse physiological functions. Simple epithelia consist of monolayered cells that form a sheet through cell-cell adherens junctions and attach to a basement membrane that lies underneath (1). Epithelial cells are polarized with the apical surface exposed, or free from cell-cell contact and the basal side lying in contact with a basement membrane or stromal cells. The cellfree apical space and basal contact are unique hallmarks of an epithelium and are likely positioning cues for the surface localization of the epithelial cells (2), although the molecular details and genes critical for epithelial cell surface positioning are yet uncertain and undefined. The concept that cues are required for epithelial surface positioning is also reinforced by observations of the disorganized growth of carcinomas. Carcinoma cells can be viewed as epithelial cells that have lost their ability to perceive surface positioning cues, and the neoplastic cells no longer obey the constraint imposed by tissue organization (3).The early embryos of vertebrates, especially mammalian species, have great plasticity, and significant cell dispersal, movement, and migration occur before their final positioning and acquisition of cell fates (4). Shown in the classical cell sorting experiments by Townes and Holtfreter (5), early embryonic amphibian cells of epidermis, endoderm, mesoderm, and neural plate, if dispersed, are able to segregate spontaneously upon aggregation, indicating cell positioning is an autonomous property.The primitive endoderm of mammalian early embryos is the first typical epithelial cell type derived that is capable of producing a basement membrane (6). Recent understanding is that the primitive endoderm cells arise from the differentiation of the pluripotent cells of the inner cell mass and migrate out to the surface to form the primitive endoderm layer (7...