Molecular Characterization of Peripherin-2 and Rom-1 Mutants Responsible for Digenic Retinitis Pigmentosa

Peripherin-2 and Rom-1 are homologous tetraspanning membrane proteins that assemble into noncovalent tetramers and higher order disulfide-linked oligomers implicated in photoreceptor disc morphogenesis. Individuals who coinherit a L185Pperipherin-2 mutation and a null or G113Erom-1 mutation are afflicted with retinitis pigmentosa, whereas individuals who inherit only one defective gene are normal. We examined the expression, subunit assembly, and disulfide-mediated oligomerization of L185P and L185A peripherin-2 and L188P Rom-1 by velocity sedimentation, co-immunoprecipitation, and cross-linking. These mutants formed noncovalent dimers under disulfide-reducing conditions but failed to assemble into core tetramers. Under nonreducing conditions, L185P dimers formed disulfide-linked tetramers but not higher order oligomers. L185P coassembled with wild-type peripherin-2 and Rom-1 to form tetramers and higher order disulfide-linked oligomers characteristic of the wild-type proteins. The G113E Rom-1 mutant expressed 20-fold lower than wild-type Rom-1, indicating that it behaves mechanistically as a null allele. We conclude that Leu185 of peripherin-2 (Leu188 of Rom-1) is critical for tetramer but not dimer formation and that the core tetramer has 2-fold symmetry. Peripherin-2-containing tetramers are required for higher order disulfide-linked oligomer formation. The level of these oligomers is critical for stable photoreceptor disc formation and the digenic retinitis pigmentosa disease phenotype. Peripherin-2 and Rom-1 are homologous tetraspanning membrane proteins that assemble into noncovalent tetramers and higher order disulfide-linked oligomers implicated in photoreceptor disc morphogenesis. Individuals who coinherit a L185Pperipherin-2 mutation and a null or G113Erom-1 mutation are afflicted with retinitis pigmentosa, whereas individuals who inherit only one defective gene are normal. We examined the expression, subunit assembly, and disulfide-mediated oligomerization of L185P and L185A peripherin-2 and L188P Rom-1 by velocity sedimentation, co-immunoprecipitation, and cross-linking. These mutants formed noncovalent dimers under disulfide-reducing conditions but failed to assemble into core tetramers. Under nonreducing conditions, L185P dimers formed disulfide-linked tetramers but not higher order oligomers. L185P coassembled with wild-type peripherin-2 and Rom-1 to form tetramers and higher order disulfide-linked oligomers characteristic of the wild-type proteins. The G113E Rom-1 mutant expressed 20-fold lower than wild-type Rom-1, indicating that it behaves mechanistically as a null allele. We conclude that Leu185 of peripherin-2 (Leu188 of Rom-1) is critical for tetramer but not dimer formation and that the core tetramer has 2-fold symmetry. Peripherin-2-containing tetramers are required for higher order disulfide-linked oligomer formation. The level of these oligomers is critical for stable photoreceptor disc formation and the digenic retinitis pigmentosa disease phenotype. wild-type retinitis pigmentosa phosphate-buffered saline dithiothreitol N-ethylmaleimide Peripherin-2 (also known as peripherin/Rds) and Rom-1 are tetraspanning membrane proteins localized along the rim region of rod and cone photoreceptor outer segment discs (1Molday R.S. Hicks D. Molday L. Invest. Ophthalmol. Vis. Sci. 1987; 28: 50-61PubMed Google Scholar, 2Connell G.J. Molday R.S. Biochemistry. 1990; 29: 4691-4698Crossref PubMed Scopus (141) Google Scholar, 3Bascom R.A. Manara S. Collins L. Molday R.S. Kalnins V.I. McInnes R.R. Neuron. 1992; 8: 1171-1184Abstract Full Text PDF PubMed Scopus (205) Google Scholar, 4Arikawa K. Molday L.L. Molday R.S. Williams D.S. J. Cell Biol. 1992; 116: 659-667Crossref PubMed Scopus (230) Google Scholar, 5Moritz O.L. Molday R.S. Invest. Ophthalmol. Vis. Sci. 1996; 37: 352-362PubMed Google Scholar). Peripherin-2 is critical to the formation and stabilization of photoreceptor outer segments, since rds mice homozygous for a null mutation in the peripherin-2 gene Prph2 fail to develop outer segments, and heterozygous rds mice form highly disorganized structures (6Sanyal S. Jansen H.G. Neurosci. Lett. 1981; 21: 23-26Crossref PubMed Scopus (166) Google Scholar, 7Hawkins R.K. Jansen H.G. Sanyal S. Exp. Eye Res. 1985; 41: 701-720Crossref PubMed Scopus (158) Google Scholar, 8Travis G.H. Brennan M.B. Danielson P.E. Kozak C.A. Sutcliffe J.G. Nature. 1989; 338: 70-73Crossref PubMed Scopus (314) Google Scholar, 9Connell G. Bascom R. Molday L. Reid D. McInnes R.R. Molday R.S. Proc. Natl. Acad. Sci. U. S. A. 1991; 88: 723-726Crossref PubMed Scopus (237) Google Scholar). In contrast, Rom-1 plays a more ancillary role in outer segment structure, since homozygous Rom-1 knockout mice develop outer segments that are only mildly altered in appearance (10Clarke G. Goldberg A.F. Vidgen D. Collins L. Ploder L. Schwarz L. Molday L.L. Rossant J. Szel A. Molday R.S. Birch D.G. McInnes R.R. Nat. Genet. 2000; 25: 67-73Crossref PubMed Scopus (131) Google Scholar). Peripherin-2 and Rom-1 share many structural properties (3Bascom R.A. Manara S. Collins L. Molday R.S. Kalnins V.I. McInnes R.R. Neuron. 1992; 8: 1171-1184Abstract Full Text PDF PubMed Scopus (205) Google Scholar, 2Connell G.J. Molday R.S. Biochemistry. 1990; 29: 4691-4698Crossref PubMed Scopus (141) Google Scholar, 11Travis G.H. Sutcliffe J.G. Bok D. Neuron. 1991; 6: 61-70Abstract Full Text PDF PubMed Scopus (242) Google Scholar). They are 35% identical in sequence and contain four putative membrane-spanning segments, a large intradiscal loop that connects the third and fourth transmembrane segments, and a relatively long C-terminal segment on the cytoplasmic side of the disc membrane (Fig.1) implicated in membrane fusion (12Boesze-Battaglia K. Lamba O.P. Napoli Jr., A.A. Sinha S. Guo Y. Biochemistry. 1998; 37: 9477-9487Crossref PubMed Scopus (73) Google Scholar). The large intradiscal loop contains seven conserved cysteine residues. One cysteine, Cys150, forms intermolecular disulfide bonds, whereas the other six cysteine residues contribute to protein folding presumably through intramolecular disulfide bonds (13Goldberg A.F. Loewen C.J. Molday R.S. Biochemistry. 1998; 37: 680-685Crossref PubMed Scopus (102) Google Scholar). Peripherin-2 and Rom-1 subunits interact noncovalently to form a mixture of homo- and heterotetrameric core complexes (14Loewen C.J. Molday R.S. J. Biol. Chem. 2000; 275: 5370-5378Abstract Full Text Full Text PDF PubMed Scopus (103) Google Scholar). Subunit assembly is mediated through interactions involving the large intradiscal loop (13Goldberg A.F. Loewen C.J. Molday R.S. Biochemistry. 1998; 37: 680-685Crossref PubMed Scopus (102) Google Scholar, 15Goldberg A.F. Molday R.S. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 13726-13730Crossref PubMed Scopus (105) Google Scholar, 16Kedzierski W. Weng J. Travis G.H. J. Biol. Chem. 1999; 274: 29181-29187Abstract Full Text Full Text PDF PubMed Scopus (31) Google Scholar). Peripherin-2-containing core complexes can link together through Cys150-mediated disulfide bonds to form higher order oligomers, which are thought to play a key role in the formation and stabilization of the hairpin rim region of outer segment discs (14Loewen C.J. Molday R.S. J. Biol. Chem. 2000; 275: 5370-5378Abstract Full Text Full Text PDF PubMed Scopus (103) Google Scholar). In support of this concept, Wrigley et al. (17Wrigley J.D. Ahmed T. Nevett C.L. Findlay J.B. J. Biol. Chem. 2000; 275: 13191-13194Abstract Full Text Full Text PDF PubMed Scopus (67) Google Scholar) have shown that in vitro expression of wild-type (WT)1 peripherin-2 under nonreducing conditions results in the flattening of microsomal vesicles, whereas expression of WT peripherin-2 under disulfide-reducing conditions or expression of several peripherin-2 mutants including the C150S mutant under nonreducing conditions produces characteristic spherical microsomal vesicles. Mutations in the Prph2 gene have been linked to a variety of human retinal degenerative diseases including autosomal dominant retinitis pigmentosa (RP), macular dystrophy, and related pattern dystrophies (18Farrar G.J. Kenna P. Jordan S.A. Kumar-Singh R. Humphries M.M. Sharp E.M. Sheils D.M. Humphries P. Nature. 1991; 354: 478-480Crossref PubMed Scopus (357) Google Scholar, 19Kajiwara K. Hahn L.B. Mukai S. Travis G.H. Berson E.L. Dryja T.P. Nature. 1991; 354: 480-483Crossref PubMed Scopus (380) Google Scholar, 20Saga M. Mashima Y. Akeo K. Oguchi Y. Kudoh J. Shimizu N. Hum. Genet. 1993; 92: 519-521Crossref PubMed Scopus (53) Google Scholar, 21Wells J. Wroblewski J. Keen J. Inglehearn C. Jubb C. Eckstein A. Jay M. Arden G. Bhattacharya S. Fitzke F. Bird A. Nat. Genet. 1993; 3: 213-218Crossref PubMed Scopus (370) Google Scholar, 22Weleber R.G. Carr R.E. Murphey W.H. Sheffield V.C. Stone E.M. Arch. Ophthalmol. 1993; 111: 1531-1542Crossref PubMed Scopus (256) Google Scholar). Most disease-causing missense mutations are located within the large intradiscal loop of peripherin-2. Heterologous expression studies have shown that some of these mutations cause misfolding of peripherin-2 and defective subunit assembly (13Goldberg A.F. Loewen C.J. Molday R.S. Biochemistry. 1998; 37: 680-685Crossref PubMed Scopus (102) Google Scholar, 15Goldberg A.F. Molday R.S. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 13726-13730Crossref PubMed Scopus (105) Google Scholar). In contrast, there is no clear example of a mutation in Rom-1 that, by itself, causes retinal degeneration (23Dryja T.P. Hahn L.B. Kajiwara K. Berson E.L. Invest. Ophthalmol. Vis. Sci. 1997; 38: 1972-1982PubMed Google Scholar). Dryja and colleagues (23Dryja T.P. Hahn L.B. Kajiwara K. Berson E.L. Invest. Ophthalmol. Vis. Sci. 1997; 38: 1972-1982PubMed Google Scholar, 24Kajiwara K. Berson E.L. Dryja T.P. Science. 1994; 264: 1604-1608Crossref PubMed Scopus (611) Google Scholar), however, identified several families with a digenic form of autosomal dominant RP linked to mutations in both peripherin-2 and rom-1. Individuals who inherited both a L185P mutation in a peripherin-2 and a null or G113E mutation in a are with whereas who inherit only one of these mutations are normal. studies have shown that the L185P peripherin-2 mutant with Rom-1 to form core heterotetrameric but WT it is to into core A.F. Molday R.S. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 13726-13730Crossref PubMed Scopus (105) Google Scholar). In order to the of digenic autosomal dominant have a of several peripherin-2 and Rom-1 that the L185P peripherin-2 mutant into dimers that can through intermolecular disulfide bonds to form tetramers but not higher order oligomers characteristic of WT peripherin-2. The L185P however, can interact with WT Rom-1 and WT peripherin-2 to form core tetramers and higher order disulfide-linked oligomers. these studies into the of the peripherin-2 and Rom-1 complexes and that contribute to autosomal dominant digenic and have been O.L. Molday R.S. Invest. Ophthalmol. Vis. Sci. 1996; 37: 352-362PubMed Google Scholar, A.F. Loewen C.J. Molday R.S. Biochemistry. 1998; 37: 680-685Crossref PubMed Scopus (102) Google Scholar, A.F. O.L. Molday R.S. Biochemistry. PubMed Scopus Google Scholar). and by 1989; PubMed Scopus Google Scholar). a the by to the the of the peripherin-2 that the of into this by with the L185P by the into with a of of and as A.F. O.L. Molday R.S. Biochemistry. PubMed Scopus Google Scholar). of the L185P mutant under conditions as for wild-type peripherin-2 A.F. Molday R.S. Biochemistry. 1996; PubMed Scopus Google Scholar). in and and for The to the properties of the The by on a by in a for on in and for in and on protein and Scholar, of and Scholar, PubMed Scopus Google Scholar, S. 1989; PubMed Scopus Google Scholar, M. R. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus (53) Google Scholar). The of the and the of the protein to the of S. 1989; PubMed Scopus Google Scholar). the for the the and the are the and in in or The the is the of the protein the of is the is the is the The by of the of the and and The of the by the of the The by the proteins the for and for the The and of under conditions of are and The by the of for the The of the the in and by the to The for the under conditions The of the the the and the PubMed Scopus Google Scholar), is the of the protein of the L185P peripherin-2 for the for protein and for the in the A.F. Molday R.S. Biochemistry. 1996; PubMed Scopus Google with and with in the or of for The with of and for on In some with in the The for and on for Peripherin-2 the on a as (14Loewen C.J. Molday R.S. J. Biol. Chem. 2000; 275: 5370-5378Abstract Full Text Full Text PDF PubMed Scopus (103) Google Scholar). peripherin-2 to in and A.F. Molday R.S. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 13726-13730Crossref PubMed Scopus (105) Google Scholar). in the for under for in a the of the and for by to for with of mixture in the or of and to or as (14Loewen C.J. Molday R.S. J. Biol. Chem. 2000; 275: 5370-5378Abstract Full Text Full Text PDF PubMed Scopus (103) Google Scholar). the proteins to a with to peripherin-2 (1Molday R.S. Hicks D. Molday L. Invest. Ophthalmol. Vis. Sci. 1987; 28: 50-61PubMed Google Scholar), Rom-1 or D. A. P. Molday R.S. Biochemistry. PubMed Scopus Google Scholar) and for by The to a fusion protein the intradiscal loop region of Rom-1 O.L. Molday R.S. Invest. Ophthalmol. Vis. Sci. 1996; 37: 352-362PubMed Google Scholar). on with for studies have shown that under conditions the of the L185P peripherin-2 mutant is than that of the WT peripherin-2 core A.F. Molday R.S. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 13726-13730Crossref PubMed Scopus (105) Google Scholar). the of this mutant to a of the L185P protein under shown in the L185P mutant a as a with a of a that is lower than the of for the WT peripherin-2 A.F. Molday R.S. Biochemistry. 1996; PubMed Scopus Google Scholar). The and of the L185P mutant by velocity in and in in of and a of no the of and not since the of L185P peripherin-2 to the rod outer segments for A.F. Molday R.S. Biochemistry. 1996; PubMed Scopus Google the L185P mutant of as the indicating that it not the for and a of for the L185P to the of S. 1989; PubMed Scopus Google Scholar). for the for protein and the for the and of for protein and the to for the L185P protein is a L185P and dimer but than the of for the peripherin-2 tetramer rod outer segments A.F. Molday R.S. Biochemistry. 1996; PubMed Scopus Google Scholar). the L185P mutant as a or a a of studies with the L185P mutant and a related The L185P mutant is by the mutant contains a C-terminal by and a mutation is in that We the expressed L185P and proteins to interact to form a stable shown in L185P and together and with under disulfide-reducing conditions, mutant in the whereas the L185P mutant only in the that expressed L185P and proteins not to form a that the the L185P mutant but not the L185P and mutants can into a with the L185P and The with to intermolecular disulfide bonds and in in the of to (14Loewen C.J. Molday R.S. J. Biol. Chem. 2000; 275: 5370-5378Abstract Full Text Full Text PDF PubMed Scopus (103) Google Scholar). The to a and the and for L185P and by that of the L185P protein to the of the protein with the L185P peripherin-2. The by velocity The that to the expressed L185P the of the that form the of peripherin-2 and core peripherin-2 is the of peripherin-2 as in peripherin-2 L188P expressed in in to and for and in a under on (13Goldberg A.F. Loewen C.J. Molday R.S. Biochemistry. 1998; 37: 680-685Crossref PubMed Scopus (102) Google is the of A.F. Molday R.S. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 13726-13730Crossref PubMed Scopus (105) Google Scholar) as in A.F. Loewen C.J. Molday R.S. Biochemistry. 1998; 37: 680-685Crossref PubMed Scopus (102) Google in a expressed in in to and for and in a under on the L185P mutant into a shown in a of L185P peripherin-2 as a dimer with that L185P is a a the and studies that the L185P peripherin-2 mutant as a dimer under disulfide-reducing the role of on the subunit assembly of the of a L185A mutant under disulfide-reducing The L185A as a characteristic of a dimer mutant coassembled with Rom-1 into a core not Rom-1 contains a The L188P Rom-1 mutant as a with of a that is than that of WT Rom-1 tetramer and with a dimer L185P peripherin-2 can assemble with L188P Rom-1, the L185P peripherin-2 and L188P Rom-1 with and the with that a of L188P Rom-1 with L185P peripherin-2. with of a a L185P and a L188P and with formation. The WT peripherin-2 core tetramer forms higher disulfide-linked oligomers under nonreducing conditions (14Loewen C.J. Molday R.S. J. Biol. Chem. 2000; 275: 5370-5378Abstract Full Text Full Text PDF PubMed Scopus (103) Google Scholar). the L185P peripherin-2 dimer forms disulfide-linked oligomers, the L185P mutant with and in in the of The to velocity sedimentation, and on nonreducing The velocity One of a to the for L185P this of on nonreducing of a that is to that for WT peripherin-2 core this disulfide-linked These results that L185P peripherin-2 core dimers can link together through intermolecular disulfide bonds to form a this not form the higher order oligomers for WT peripherin-2 studies have shown that both WT and L185P peripherin-2 assemble with WT Rom-1 to form heterotetrameric complexes under conditions A.F. Molday R.S. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 13726-13730Crossref PubMed Scopus (105) Google Scholar). these complexes can form higher order disulfide-linked oligomers, examined the of WT and the L185P under nonreducing shown in the WT as of the as a in of intermolecular disulfide bonds, and as oligomer presumably a of intermolecular disulfide The of L185P is shown in a for of the and of a for of the and both and and oligomer for of and disulfide-linked These results that of the L185P peripherin-2 mutant with Rom-1 into higher order disulfide-linked complexes to that for WT Individuals with autosomal dominant RP both WT and mutant peripherin-2 L185P peripherin-2 mutant with WT the L185P mutant with peripherin-2 the C-terminal and the mutation that that under disulfide-reducing conditions, of the with the L185P peripherin-2 of this a of and Under nonreducing conditions, the velocity of L185P the of higher order disulfide-linked oligomers as as and In the expressed WT the as WT peripherin-2 indicating that the of the no on subunit of individuals with autosomal dominant digenic RP have a G113Erom-1 missense mutation in to the L185Pperipherin-2 mutation (23Dryja T.P. Hahn L.B. Kajiwara K. Berson E.L. Invest. Ophthalmol. Vis. Sci. 1997; 38: 1972-1982PubMed Google Scholar). We the protein expression level of G113E Rom-1 with that of WT Rom-1 in In four the level of G113E Rom-1 expression that of WT that G113E Rom-1 expression to this digenic studies have shown that the L185P peripherin-2 mutant expressed in has a that is than that of the WT peripherin-2 core tetramer A.F. Molday R.S. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 13726-13730Crossref PubMed Scopus (105) Google Scholar). We have the of this mutant by and cross-linking. a that a and both and that the L185P mutant as a dimer under disulfide-reducing The for the of L185P dimer by is with the of the of the this A.F. Molday R.S. Biochemistry. 1996; PubMed Scopus Google Scholar). in can in the that the protein the of in and S. 1989; PubMed Scopus Google Scholar). The of the protein can contribute to for the of the in the of are for proteins. The however, is not only on the of the protein but on in the of a as as the of of in the of membrane proteins S. 1989; PubMed Scopus Google Scholar). WT and L185P peripherin-2 are known to in not The of the however, is not known and not into the The of the to the however, is to since studies that peripherin-2 is not G.J. Molday R.S. Biochemistry. 1990; 29: 4691-4698Crossref PubMed Scopus (141) Google Scholar, A.F. O.L. Molday R.S. Biochemistry. PubMed Scopus Google Scholar). The peripherin-2 mutant contain a of in the which can have a on the The of in this of the of the peripherin-2 which in the for the of the peripherin-2 The role of the conserved in subunit assembly by several The L185A peripherin-2 mutant the L185P formed a core dimer expressed but into a with that defective formation of the L185P mutant is to the of a and not the of a The of these mutants to interact with Rom-1 to form a tetramer that the mutants are not Rom-1 has a within a conserved segment of the large intradiscal loop of this with Rom-1 into these studies that peripherin-2 and in Rom-1 play in the of dimers into core tetramers. The that these mutants as dimers and not into the of the peripherin-2 and Rom-1 core The interactions for dimer formation are for tetramer since is not on in peripherin-2 in this that the WT peripherin-2 and Rom-1 core tetramer symmetry. to a in which the core tetramer can as a of The for dimer formation is not known the the of L185P peripherin-2 to with L188P Rom-1 to form a that the of these the The of and other residues to the formation and of interactions is for proteins of the J.D. T. Science. 1991; PubMed Scopus Google the subunit of Y. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google proteins and R.A. Biol. 1997; PubMed Scopus (67) Google and membrane proteins D.M. Science. 1997; PubMed Scopus Google Scholar) and N. Y. Y. T. M. T. S. K. Nature. 2000; PubMed Scopus Google the of the region of the a has been N. Y. Y. T. M. T. S. K. Nature. 2000; PubMed Scopus Google Scholar). In this four conserved and one contribute to the dimer In the intradiscal loop region of peripherin-2 and Rom-1, there are conserved and several conserved residues in to is that some of these residues contribute to subunit interactions of peripherin-2 and many of these residues are conserved in other of the of proteins including and 1994; Full Text PDF PubMed Scopus Google for the conserved to the that these proteins as complexes subunit interactions are mediated by these residues. Peripherin-2-containing core tetramers link together through Cys150-mediated intermolecular disulfide bonds to form higher order oligomers (14Loewen C.J. Molday R.S. J. Biol. Chem. 2000; 275: 5370-5378Abstract Full Text Full Text PDF PubMed Scopus (103) Google Scholar). In this have shown that L185P dimers link together to form a disulfide-linked tetramer but this not assemble into the higher order oligomers for WT peripherin-2. that core tetramer formation is not required for intermolecular disulfide but it is required for the of higher order oligomers. together with studies that C150S peripherin-2 mutants form core noncovalent but not higher order oligomers (14Loewen C.J. Molday R.S. J. Biol. Chem. 2000; 275: 5370-5378Abstract Full Text Full Text PDF PubMed Scopus (103) Google Scholar), that higher order oligomerization both noncovalent and The noncovalent interactions in higher order oligomerization are to to in of Cys150-mediated intermolecular disulfide bonds tetramers a within the intradiscal loop that noncovalent interactions for oligomers to in outer segment disc formation. In vitro expression of WT peripherin-2 under nonreducing conditions has been to in microsomal vesicles, whereas expression of WT peripherin-2 under conditions or expression of C150S peripherin-2 under nonreducing conditions results in (17Wrigley J.D. Ahmed T. Nevett C.L. Findlay J.B. J. Biol. Chem. 2000; 275: 13191-13194Abstract Full Text Full Text PDF PubMed Scopus (67) Google Scholar). is not known the disulfide-linked L185P tetramers cause the flattening of these or higher order oligomers are required for this Individuals with autosomal dominant digenic RP a along with the mutant allele. in more the for this examined the of L185P peripherin-2 mutant with WT peripherin-2 and WT of L185P and WT peripherin-2 in a mixture of and core complexes that through intermolecular disulfide bonds to form oligomers. L185P peripherin-2 with Rom-1 to form core tetramers as shown A.F. Molday R.S. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 13726-13730Crossref PubMed Scopus (105) Google Scholar). the WT the L185P intermolecular disulfide bonds into disulfide-linked oligomers, the of these conclude that the L185P mutant can interact with WT proteins to form higher order oligomers that contribute to the formation and stabilization of outer segment Individuals with autosomal dominant digenic RP linked to the L185P mutation in peripherin-2 coinherit a mutation In families to a or of the in of the and in a null K. Berson E.L. Dryja T.P. Science. 1994; 264: 1604-1608Crossref PubMed Scopus (611) Google Scholar). Dryja et al. (23Dryja T.P. Hahn L.B. Kajiwara K. Berson E.L. Invest. Ophthalmol. Vis. Sci. 1997; 38: 1972-1982PubMed Google Scholar) identified a with digenic RP that has a G113E mutation in along with the L185P mutation in peripherin-2. the G113E missense mutation in the third putative transmembrane segment of Rom-1 that of a into a membrane-spanning segment the stable expression of results that the level of G113E Rom-1 expression in is 20-fold lower than WT to the level of G113E Rom-1 expression, to the properties of this mutant or with peripherin-2. the of the expression level of this it is to conclude that the G113E mutant in Rom-1 behaves mechanistically as a null dominant digenic RP is a disease the of a L185P peripherin-2 and null A.F. Molday R.S. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 13726-13730Crossref PubMed Scopus (105) Google Scholar, T.P. Hahn L.B. Kajiwara K. Berson E.L. Invest. Ophthalmol. Vis. Sci. 1997; 38: 1972-1982PubMed Google Scholar, 24Kajiwara K. Berson E.L. Dryja T.P. Science. 1994; 264: 1604-1608Crossref PubMed Scopus (611) Google Scholar). Individuals who inherit only one of these mutant are normal. that the level of oligomers is critical in the of this the L185P peripherin-2 mutant is of and higher order disulfide-linked oligomers, individuals who inherit this mutation along with a null in Rom-1 have a level of oligomers the critical level required for the formation of stable outer The of outer segments to photoreceptor degeneration as for heterozygous R.K. Jansen H.G. Sanyal S. Exp. Eye Res. 1985; 41: 701-720Crossref PubMed Scopus (158) Google Scholar). In the of individuals who inherit only the L185Pperipherin-2 a of the L185P assemble with WT Rom-1 as as WT peripherin-2 to form oligomers. have the of the level of oligomers the to form stable outer These individuals photoreceptor degeneration and have Individuals who inherit only a null to have of and the level of of Rom-1, however, have no on outer segment or photoreceptor as shown in studies of heterozygous knockout mice (10Clarke G. Goldberg A.F. Vidgen D. Collins L. Ploder L. Schwarz L. Molday L.L. Rossant J. Szel A. Molday R.S. Birch D.G. McInnes R.R. Nat. Genet. 2000; 25: 67-73Crossref PubMed Scopus (131) Google Scholar). individuals who inherit only a null or a G113E missense mutation in have a level of oligomers to support photoreceptor outer segment and photoreceptor The L185P peripherin-2 mutant is peripherin-2 mutants that cause autosomal dominant the and peripherin-2 mutants are highly and not form core tetramers or higher order oligomers in the or of Rom-1 (13Goldberg A.F. Loewen C.J. Molday R.S. Biochemistry. 1998; 37: 680-685Crossref PubMed Scopus (102) Google Scholar, C.J. Molday R.S. J. Biol. Chem. 2000; 275: 5370-5378Abstract Full Text Full Text PDF PubMed Scopus (103) Google Scholar). expression of the mutant to in the formation of (17Wrigley J.D. Ahmed T. Nevett C.L. Findlay J.B. J. Biol. Chem. 2000; 275: 13191-13194Abstract Full Text Full Text PDF PubMed Scopus (67) Google Scholar). individuals with these mutations have a level of oligomers in the or of Rom-1, in disorganized outer segment discs and photoreceptor dominant of these however, contribute to photoreceptor degeneration and the autosomal dominant RP phenotype. We Goldberg for the

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