Molecular characterization of photoreceptor peripherin-2 and rom-1 complexes and role in disc morphogenesis and retinal degeneration

Peripherin-2 and rom-1 are homologous integral membrane proteins of the tetraspanin superfamily that form multisubunit complexes in the rims of photoreceptor outer segment discs. Peripherin-2 is critical for formation and maintenance of rod and cone discs, while rom-1 is involved in regulation of this process. Mutations in peripherin-2 cause various forms of human retinal degeneration including retinitis pigmentosa (RP) and macular degeneration (MD). Peripherin-2 and rom-1 contain seven highly conserved cysteines in the intradiscal loop region, two of which in peripherin-2 are linked to RP. Each of these cysteines in peripherin-2 has been individually replaced by serine to determine its role in folding and subunit assembly. Six of the seven conserved cysteine residues in the intradiscal loop of peripherin-2 are essential for proper core tetramer formation with rom-1. They likely constitute three intramolecular disulfide bonds crucial to the proper folding of the subunits. Mutations at C165 and C214 cause RP. The remaining cysteine, C150, is not involved in tetramer formation, but is solely responsible for disulfide-mediated oligomerization of tetramers into higher order complexes. Core tetramer formation is required for disulfide-mediated oligomerization and for targeting to rod outer segments. RP-causing mutations in peripherin-2 prevent tetramer and higher-order oligomer formation and cause mistargeting to rod inner segments.Disulfide-mediated oligomerization plays a role in disc rim and incisure formation and involves both noncovalent and covalent (disulfide bonds) interactions. Rom-1 functions in the regulation of disulfide-mediated oligomerization by inhibiting disulfide-linking of tetramers. Human RP caused by mutations in peripherin-2 results from (1) having decreased levels of protein in the outer segments, and (2) by interference with the function of WT protein in outer segments in a dominant-negative manner.Medicine, Faculty ofBiochemistry and Molecular Biology, Department ofGraduat