The tetraspanin disc proteins, peripherin-2 and ROM1, facilitate CNG channel localization to the rod outer segment

The light-responsive outer segment of rod photoreceptors is composed of two distinct membrane subdomains: discs and the plasma membrane that surrounds them. To understand how proteins are delivered to these subdomains, we focused on the plasma membrane-specific cyclic nucleotide-gated (CNG) channel. It was previously reported that the {beta}1 subunit of the CNG channel is absent from rods in peripherin-2 knockout (Rds-/-) mice. It is unclear how the disc-resident protein peripherin-2 would impact the localization of CNG{beta}1 in the plasma membrane. In this study, we investigate how peripherin-2 is engaged in CNG channel delivery to the outer segment. We overexpressed a MYC-tagged CNG{beta}1 in Rds-/- rods and found that it is trapped in intracellular membranes. CNG{beta}1 localization was restored to the outer segment when full-length, FLAG-tagged peripherin-2 was expressed. This led us to investigate the region of peripherin-2 that is required for CNG{beta}1 delivery by expressing peripherin-2 chimeras containing either the N-terminus, tetraspanin core, or C-terminus. We show that the peripherin-2 tetraspanin domain is sufficient to localize CNG{beta}1 to the outer segment. Recent studies have shown that peripherin-2 and ROM1 act redundantly in outer segment disc formation and enclosure. Consistent with this, we found that expression of ROM1 chimeras containing its tetraspanin region could also restore CNG{beta}1 localization. We postulated that the structural properties of these tetraspanin proteins could facilitate CNG channel delivery to the outer segment. Interestingly, this notion is supported by endogenous staining of CNG{beta}1, which reappears in aged Rds-/- rods that have produced ciliary membrane protrusions.