USH1 and USH2 proteins in the calyceal processes of adult amphibian photoreceptors. (A) Adult Xenopus laevis photoreceptors. Multiple indentations run along the photoreceptor outer segment (OS) from its base to its tip. These indentations are occupied at their base by well-developed calyceal processes (CP). (B) Co-immunoprecipitation of USH1 proteins. Using adult Xenopus retinal extracts, the anti-sans antibodies anti-S1SAM and anti-S1Ank, but not protein G alone, immunoprecipitate endogenous myosin VIIa (top panels) and harmonin (bottom panels). (C and D) Flat-mount sections of Xenopus and bullfrog photoreceptors. The USH1 proteins are detected nearby the F-actin–labeled (red) calyceal processes (C), whereas the USH2 protein Vlgr1 is restricted to the periciliary ridge complex (PRC) region surrounding the labeled connecting cilium (blue immunolabeling, highlighted on the scanning electron micrograph of a broken rod). (E) Retinas from adult Xenopus (n = 3) were incubated in Ringer solution alone, or containing subtilisin, or subtilisin followed by BAPTA. Subtilisin treatment did not affect the attachment of the calyceal processes to the outer segment, but the addition of BAPTA resulted in the detachment of the calyceal processes, which lost their connection to the plasma membrane of the outer segment, sometimes along their entire length (arrows). Bars: (A and E) 1 µm; (C and D) 2 µm.