ROM-1 potentiates photoreceptor specific membrane fusion processes.
Photoreceptor outer segment (OS) renewal requires a series of tightly regulated membrane fusion events which are mediated by a fusion complex containing protein and lipid components. The best characterized of these components, is a unique photoreceptor specific tetraspanin, peripherin/rds (P/rds, a.k.a., peripherin-2, Rds and Prph). In these studies we investigated the role of peripherin's non-glycosylated homolog, ROM-1, in OS fusion using a COS cell heterologous expression system and a well characterized cell free fusion assay system. Membranes isolated from COS-7 cells transfected with either FLAG-tagged P/rds or HA-tagged ROM-1 or both proteins were assayed for their ability to merge with fluorescently labeled OS plasma membrane (PM). Such membrane merger is one measure of membrane fusogenicity. The highest percent fusion was observed when the proteins were co-expressed. Furthermore detailed analysis of the fusion kinetics between fluorescently labeled PM and proteo-liposomes containing either, pure P/rds, pure ROM-1 or the ROM-1-P/rds complex clearly demonstrated that optimal fusion requires an ROM-1/P/rds complex. Proteo-liposomes composed of ROM-1 alone were not fusogenic. Peptide competition studies suggest that optimization of fusion may be due to the formation of a fusion competent peripherin/rds C-terminus in the presence of ROM-1. These studies provide further support for the hypothesis that a P/rds dependent membrane fusion complex is involved in photoreceptor renewal processes.
Boesze-Battaglia
Kathleen
K
Department of Biochemistry, School of Dental Medicine, University of Pennsylvania, 240 South 40th Street, Philadelphia, PA 19104, USA. battagli@biochem.dental.upenn.edu
Stefano
Frank P
FP
Fitzgerald
Catherine
C
Muller-Weeks
Susan
S
D000818:Animals; D015153:Blotting, Western; D019556:COS Cells; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D002474:Cell-Free System; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D005136:Eye Proteins / Q000378:metabolism / Q000502:physiology*; D008081:Liposomes; D008561:Membrane Fusion / Q000502:physiology*; D008565:Membrane Proteins / Q000302:isolation & purification / Q000378:metabolism / Q000502:physiology*; D012374:Rod Cell Outer Segment / Q000502:physiology*; D014162:Transfection
D005136:Eye Proteins; D008081:Liposomes; D008565:Membrane Proteins