Figure 2

Schematic diagrams of HRP-rhodopsin reporter, its expected expression in different disc genesis models, and its expression pattern in transfected rods. (A) A diagram of the HRP-rhodopsin reporter in the context of its transmembrane helices. The N-terminus is tagged with HRP and the signal sequence (SS) of Drosophila wingless gene, while the C-terminus is tagged with GFP and the rhodopsin 1D4 epitope. (B) A schematic diagram delineating the predicted distribution of DAB precipitates (burgundy) in the scenario that the OS base has evaginated PM protruding into the extracellular (Ec) space. Inset depicts the DAB deposits that would be expected on the convex side of evaginated PM outfolds that are filled by cytosol (Cy; gray). (C) A schematic diagram delineating the predicted distribution of DAB precipitates (burgundy) in the scenario that the base of the OS is completely enveloped by the PM and surrounded by cytosol. DAB would also be observed on the luminal sides of rhodopsin-bearing vesicles and on the luminal (Lu) side of disc membranes (inset). (D) A representative confocal image of a retinal slice containing HRP-rhodopsin transfected rods. (E) An electron micrograph revealing the OS of three rods after DAB reactions. Cell No. 1 is positive, whereas No. 2 and No. 3 are negative for HRP activity. An arrow points to the disc incisures of cell No. 2.