Conservative amino acid substitutions are tolerated at T4 but not at T17. Transgenic animals expressing hWT, hT4I, hT17M, or hT17V rhodopsins (n = 20 per group) were generated and raised in cyclic light. A, Plot of transgenic opsin expression levels versus total rod opsin levels derived from solubilized transgenic eye extracts. B, C, Confocal micrographs of cryosections from transgenic retinas stained with WGA (B) or labeled with mAb 2B2 (green) and counterstained with WGA (red) and Hoescht nuclear dye (blue; C). Expression of hT17V rhodopsin resulted in loss of rod OS, whereas expression of hT4I did not affect rod density or morphology (B). Both T4I and T17V rhodopsins localized predominantly to the rod OS (C). rpe, Retinal pigment epithelium; ros, rod outer segment; ris, rod inner segment; onl, outer nuclear layer. Scale bars: B, 20 μm; C, 5 μm.