Purification of a membrane protein distributed in a topographic gradient in chicken retina

Antigenic molecules termed TOP, which are distributed in a dorsal > ventral concentration gradient in chicken retina, are expressed early in development (by 48 hr after fertilization) in the optic cup of chicken embryos and continue to be expressed in retina thereafter. ³⁵S-labeled-TOP-antibody complexes were purified by protein A-Sepharose column chromatography and subjected to NaDodDO₄/polyacrylamide gel electrophoresis and autoradiography. TOP also was purified from dorsal retina by anti-TOP IgG-Affi-Gel 10 affinity dolumn chromatography. In both cases, one major band of protein at M(r) ≃ 47,000 was obtained. A protein of M(r) ≃ 47,000 also was purified from chicken embryo brain. Cultured cells dissociated from 8-day chicken embryo retinas accumulated the amount of TOP expected of cells in the intact retina, depending on the position of the cells in the retina. TOP accumulations by cells dissociated from dorsal or ventral retina, mixed in different proportions, and cocultured were additive. These results show that TOP is a protein, that the gradient of TOP is established early in development, and that perpetuation of the gradient does not depend on the continuous presence of an extracellular gradient of diffusable molecules or on maintenance of interactions between cells.