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Shannon Conley, Michael W Stuck, Rahel Zulliger, Muna I Naash; Complex size and composition impact RDS/ROM-1 trafficking pathway. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):4640.
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© ARVO (1962-2015); The Authors (2016-present)
Peripherin-2 (also known as retinal degeneration slow, RDS) is required for photoreceptor outer segment (OS) rim formation, and RDS mutations cause widely varying retinal diseases. Recent evidence has suggested that RDS can traffic to the OS via an unconventional secretory pathway which bypasses the trans-Golgi. To function, RDS forms multiple types of complexes with its non-glycosylated homologue rod outer segment membrane protein-1 (ROM-1), and our goal was to understand to what extent unconventional trafficking is affected by RDS complex formation and by pathogenic RDS mutations.
Glycosylated proteins which have bypassed the trans-Golgi (i.e. unconventionally secreted) do not become resistant to endoglycosidase H (EndoH). Thus retinal extracts from wild-type (WT), rom1-/-, and transgenic/knock-in mice carrying mutant RDS underwent glycosidase analysis, sometimes coupled with immunoprecipitation or velocity sedimentation to separate RDS/ROM-1 complexes.
In the WT murine retina, ~60% of RDS was EndoH sensitive. The fraction of RDS which remained sensitive to EndoH correlated with complex size: large RDS oligomers were almost completely EndoH sensitive, while ~60% of RDS found in octamers was EndoH sensitive, and only ~10% of RDS found in tetramers was EndoH sensitive. In the rom1-/- retinas virtually all RDS became EndoH sensitive. RDS mutations altered EndoH sensitivity consistent with their previously known effects on complex formation. For example, Y141C-RDS (which targets to the OS but forms abnormally large RDS oligomers) was more sensitive to EndoH than WT, while C150S-RDS (which forms tetramers but not larger oligomers) was less sensitive to EndoH than WT RDS.
These results suggest that in mice, RDS traffics by both conventional and unconventional pathways, and that both complex size and ROM-1 can significantly influence the trafficking pathway selected. These data indicate that RDS complex formation is determined early, prior to arrival in the OS, and that mutations which do not alter gross OS targeting of RDS may still cause subtle alterations in the trafficking pathway with as yet unknown consequences. Given their preservation across species, the different trafficking pathways they use to get to the OS, and their alterations in the presence of pathogenic mutations, determining the functional role of the different types of RDS/ROM-1 complexes is critical.
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