scholarly journals Rab28 is localised to photoreceptor outer segments, regulates outer segment shedding but is not linked to retinal degeneration in zebrafish

2019 ◽  
Author(s):  
Stephen P. Carter ◽  
Ailís L. Moran ◽  
David Matallanas ◽  
Gavin J. McManus ◽  
Oliver E. Blacque ◽  
...  
Keyword(s):  
Retinal Degeneration ◽  
Outer Segment ◽  
Membrane Surface ◽  
Photoreceptor Outer Segment ◽  
Membrane Surface Area ◽  
Photoreceptor Outer Segments ◽  
Outer Segments ◽  
Retinal Structure ◽  
Phototransduction Cascade ◽  
Shed Light

AbstractThe photoreceptor outer segment is the canonical example of a modified and highly specialised cilium, with an expanded membrane surface area in the form of discs or lamellae for efficient light detection. Many ciliary proteins are essential for normal photoreceptor function and cilium dysfunction often results in retinal degeneration leading to impaired vision. Herein, we investigate the function and localisation of the ciliary G-protein RAB28 in zebrafish cone photoreceptors. CRISPR-Cas9 generated rab28 mutant zebrafish display a reduction in shed outer segment material in the RPE at 1 month post fertilisation (mpf), but otherwise normal retinal structure and visual function up to 12 mpf. Cone photoreceptor-specific transgenic reporter lines show Rab28 localises almost exclusively to outer segments, independently of nucleotide binding. Co-immunoprecipitation analysis demonstrates tagged Rab28 interacts with components of the phototransduction cascade, including opsins, Phosphodiesterase 6C and Guanylate Cyclase 2D. Our data shed light on RAB28 function in cones and provide a model for RAB28-associated cone-rod dystrophy.

PRCD is Concentrated at the Base of Photoreceptor Outer Segments and is Involved in Outer Segment Disc Formation

2019 ◽  
Author(s):  
Gilad Allon ◽  
Irit Mann ◽  
Lital Remez ◽  
Elisabeth Sehn ◽  
Leah Rizel ◽  
...  
Keyword(s):  
Retinal Degeneration ◽  
Knockout Mice ◽  
Outer Segment ◽  
Mouse Retina ◽  
Cone Photoreceptors ◽  
Photoreceptor Outer Segment ◽  
Photoreceptor Outer Segments ◽  
Outer Segments ◽  
Rod And Cone Photoreceptors ◽  
Disc Formation

Abstract Mutations of the PRCD gene are associated with rod-cone degeneration in both dogs and humans. Prcd is expressed in the mouse eye as early as embryonic day 14. In the adult mouse retina PRCD is expressed in the outer segments of both rod and cone photoreceptors. Immunoelectron microscopy revealed that PRCD is located at the outer segment rim, and that it is highly concentrated at the base of the outer segment. Prcd-knockout mice present with progressive retinal degeneration, starting at 20 weeks of age and onwards. This process is reflected by a significant and progressive reduction of both scotopic and photopic electroretinographic responses, and by thinning of the retina, and specifically of the outer nuclear layer, indicating photoreceptor loss. Electron microscopy revealed severe damage to photoreceptor outer segments, which is associated with immigration of microglia cells to the Prcd-knockout retina, and accumulation of vesicles in the inter-photoreceptor space. Phagocytosis of photoreceptor outer segment discs by the retinal pigmented epithelium is severely reduced. Our data show that Prcd-knockout mice serve as a good model for retinal degeneration caused by PRCD mutations in humans. Our findings in these mice support the involvement of PRCD in outer segment disc formation of both rod and cone photoreceptors. Furthermore, they suggest a feedback mechanism which coordinates the rate of photoreceptor outer segment disc formation, shedding and phagocytosis. This study has important implications for understanding the function of PRCD in the retina, as well as for future development of treatment modalities for PRCD-deficiency in humans.

scholarly journals TULP1 and TUB Are Required for Specific Localization of PRCD to Photoreceptor Outer Segments

2020 ◽  
Vol 21 (22) ◽  
pp. 8677
Author(s):  
Lital Remez ◽  
Ben Cohen ◽  
Mariela J. Nevet ◽  
Leah Rizel ◽  
Tamar Ben-Yosef
Keyword(s):  
Protein Interactions ◽  
Mammalian Cells ◽  
Outer Segment ◽  
Protein Protein Interactions ◽  
Photoreceptor Outer Segment ◽  
Photoreceptor Outer Segments ◽  
Outer Segments ◽  
Yeast Two Hybrid System ◽  
Specific Localization ◽  
Recruitment System

Photoreceptor disc component (PRCD) is a small protein which is exclusively localized to photoreceptor outer segments, and is involved in the formation of photoreceptor outer segment discs. Mutations in PRCD are associated with retinal degeneration in humans, mice, and dogs. The purpose of this work was to identify PRCD-binding proteins in the retina. PRCD protein-protein interactions were identified when implementing the Ras recruitment system (RRS), a cytoplasmic-based yeast two-hybrid system, on a bovine retina cDNA library. An interaction between PRCD and tubby-like protein 1 (TULP1) was identified. Co-immunoprecipitation in transfected mammalian cells confirmed that PRCD interacts with TULP1, as well as with its homolog, TUB. These interactions were mediated by TULP1 and TUB highly conserved C-terminal tubby domain. PRCD localization was altered in the retinas of TULP1- and TUB-deficient mice. These results show that TULP1 and TUB, which are involved in the vesicular trafficking of several photoreceptor proteins from the inner segment to the outer segment, are also required for PRCD exclusive localization to photoreceptor outer segment discs.

scholarly journals Localization of peripherin/rds in the disk membranes of cone and rod photoreceptors: relationship to disk membrane morphogenesis and retinal degeneration.

1992 ◽  
Vol 116 (3) ◽  
pp. 659-667 ◽  
Author(s):  
K Arikawa ◽  
L L Molday ◽  
R S Molday ◽  
D S Williams
Keyword(s):  
Plasma Membrane ◽  
Retinal Degeneration ◽  
Outer Segment ◽  
Photoreceptor Cells ◽  
Growth Stages ◽  
Rod Outer Segments ◽  
Rod Photoreceptor ◽  
Outer Segments ◽  
Disk Membrane ◽  
Membrane Morphogenesis

The outer segments of vertebrate rod photoreceptor cells consist of an ordered stack of membrane disks, which, except for a few nascent disks at the base of the outer segment, is surrounded by a separate plasma membrane. Previous studies indicate that the protein, peripherin or peripherin/rds, is localized along the rim of mature disks of rod outer segments. A mutation in the gene for this protein has been reported to be responsible for retinal degeneration in the rds mouse. In the present study, we have shown by immunogold labeling of rat and ground squirrel retinas that peripherin/rds is present in the disk rims of cone outer segments as well as rod outer segments. Additionally, in the basal regions of rod and cone outer segments, where disk morphogenesis occurs, we have found that the distribution of peripherin/rds is restricted to a region that is adjacent to the cilium. Extension of its distribution from the cilium coincides with the formation of the disk rim. These results support the model of disk membrane morphogenesis that predicts rim formation to be a second stage of growth, after the first stage in which the ciliary plasma membrane evaginates to form open nascent disks. The results also indicate how the proteins of the outer segment plasma membrane and the disk membranes are sorted into their separate domains: different sets of proteins may be incorporated into membrane outgrowths during different growth stages of disk morphogenesis. Finally, the presence of peripherin/rds protein in both cone and rod outer segment disks, together with the phenotype of the rds mouse, which is characterized by the failure of both rod and cone outer segment formation, suggest that the same rds gene is expressed in both types of photoreceptor cells.

scholarly journals ROM1 contributes to phenotypic heterogeneity in PRPH2-associated retinal disease

2020 ◽  
Vol 29 (16) ◽  
pp. 2708-2722
Author(s):  
Daniel Strayve ◽  
Mustafa S Makia ◽  
Mashal Kakakhel ◽  
Haarthi Sakthivel ◽  
Shannon M Conley ◽  
...  
Keyword(s):  
Retinal Degeneration ◽  
Phenotypic Variability ◽  
Retinal Disease ◽  
Phenotypic Heterogeneity ◽  
Transgenic Mouse Models ◽  
Photoreceptor Outer Segments ◽  
Binding Partner ◽  
Biochemical Effects ◽  
Cone Function ◽  
Retinal Structure

Abstract Peripherin 2 (PRPH2) is a retina-specific tetraspanin protein essential for the formation of rod and cone photoreceptor outer segments (OS). Patients with mutations in PRPH2 exhibit severe retinal degeneration characterized by vast inter- and intra-familial phenotypic heterogeneity. To help understand contributors to this within-mutation disease variability, we asked whether the PRPH2 binding partner rod OS membrane protein 1 (ROM1) could serve as a phenotypic modifier. We utilized knockin and transgenic mouse models to evaluate the structural, functional and biochemical effects of eliminating one allele of Rom1 (Rom1+/−) in three different Prph2 models which mimic human disease: C213Y Prph2 (Prph2C/+), K153Del Prph2 (Prph2K/+) and R172W (Prph2R172W). Reducing Rom1 in the absence of Prph2 mutations (Rom1+/−) had no effect on retinal structure or function. However, the effects of reducing Rom1 in the presence of Prph2 mutations were highly variable. Prph2K/+/Rom1+/− mice had improved rod and cone function compared with Prph2K/+ as well as amelioration of K153Del-associated defects in PRPH2/ROM1 oligomerization. In contrast, Prph2R172W/Rom1+/− animals had worsened rod and cone function and exacerbated retinal degeneration compared with Prph2R172W animals. Removing one allele of Rom1 had no effect in Prph2C/+. Combined, our findings support a role for non-pathogenic ROM1 null variants in contributing to phenotypic variability in mutant PRPH2-associated retinal degeneration. Since the effects of Rom1 reduction are variable, our data suggest that this contribution is specific to the type of Prph2 mutation.

scholarly journals A new mouse model for retinal degeneration due to Fam161a deficiency

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Avigail Beryozkin ◽  
Chen Matsevich ◽  
Alexey Obolensky ◽  
Corinne Kostic ◽  
Yvan Arsenijevic ◽  
...  
Keyword(s):  
Mouse Model ◽  
Retinal Degeneration ◽  
Outer Segment ◽  
Ganglion Cells ◽  
Photoreceptor Outer Segment ◽  
Tem Analysis ◽  
Retinal Degenerations ◽  
Progressive Degeneration ◽  
Transmission Electron ◽  
Lacz Staining

AbstractFAM161A mutations are the most common cause of inherited retinal degenerations in Israel. We generated a knockout (KO) mouse model, Fam161atm1b/tm1b, lacking the major exon #3 which was replaced by a construct that include LacZ under the expression of the Fam161a promoter. LacZ staining was evident in ganglion cells, inner and outer nuclear layers and inner and outer-segments of photoreceptors in KO mice. No immunofluorescence staining of Fam161a was evident in the KO retina. Visual acuity and electroretinographic (ERG) responses showed a gradual decrease between the ages of 1 and 8 months. Optical coherence tomography (OCT) showed thinning of the whole retina. Hypoautofluorescence and hyperautofluorescence pigments was observed in retinas of older mice. Histological analysis revealed a progressive degeneration of photoreceptors along time and high-resolution transmission electron microscopy (TEM) analysis showed that photoreceptor outer segment disks were disorganized in a perpendicular orientation and outer segment base was wider and shorter than in WT mice. Molecular degenerative markers, such as microglia and CALPAIN-2, appear already in a 1-month old KO retina. These results indicate that a homozygous Fam161a frameshift mutation affects retinal function and causes retinal degeneration. This model will be used for gene therapy treatment in the future.

scholarly journals Kif17 phosphorylation regulates its ciliary localization and photoreceptor outer segment turnover

2018 ◽  
Author(s):  
Tylor R. Lewis ◽  
Sean R. Kundinger ◽  
Brian A. Link ◽  
Christine Insinna ◽  
Joseph C. Besharse
Keyword(s):  
Mammalian Cells ◽  
Outer Segment ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Consensus Sequence ◽  
Intraflagellar Transport ◽  
Photoreceptor Outer Segment ◽  
Transgenic Expression ◽  
Outer Segments ◽  
Ciliary Localization

AbstractBackgroundKIF17, a kinesin-2 motor that functions in intraflagellar transport, can regulate the onset of photoreceptor outer segment development. However, the function of KIF17 in a mature photoreceptor remains unclear. Additionally, the ciliary localization of KIF17 is regulated by a C-terminal consensus sequence (KRKK) that is immediately adjacent to a conserved residue (mouse S1029/zebrafish S815) previously shown to be phosphorylated by CaMKII. Yet, whether this phosphorylation can regulate the localization, and thus function, of KIF17 in ciliary photoreceptors remains unknown.ResultsUsing transgenic expression in both mammalian cells and zebrafish photoreceptors, we show that phospho-mimetic KIF17 has enhanced localization to cilia. Importantly, expression of phospho-mimetic KIF17 is associated with greatly enhanced turnover of the photoreceptor outer segment through disc shedding in a cell-autonomous manner, while genetic mutants of kif17 in zebrafish and mice have diminished disc shedding. Lastly, cone expression of constitutively active tCaMKII leads to a kif17-dependent increase in disc shedding.ConclusionsTaken together, our data support a model in which phosphorylation of KIF17 promotes its ciliary localization. In cone photoreceptor outer segments, this promotes disc shedding, a process essential for photoreceptor maintenance and homeostasis. While disc shedding has been predominantly studied in the context of the mechanisms underlying phagocytosis of outer segments by the retinal pigment epithelium, this work implicates photoreceptor-derived signaling in the underlying mechanisms of disc shedding.

scholarly journals Distribution of guanylate cyclase within photoreceptor outer segments

1996 ◽  
Vol 109 (7) ◽  
pp. 1803-1812
Author(s):  
M.A. Hallett ◽  
J.L. Delaat ◽  
K. Arikawa ◽  
C.L. Schlamp ◽  
F. Kong ◽  
...  
Keyword(s):  
Guanylate Cyclase ◽  
Actin Filaments ◽  
Outer Segment ◽  
Essential Role ◽  
Photoreceptor Cells ◽  
Rod Photoreceptor ◽  
Light Activation ◽  
Photoreceptor Outer Segment ◽  
Photoreceptor Outer Segments ◽  
Vertebrate Photoreceptor

Guanylate cyclases play an essential role in the recovery of vertebrate photoreceptor cells after light activation. Here, we have investigated how one such guanylate cyclase, RetGC-1, is distributed within light- and dark-adapted rod photoreceptor cells. Guanylate cyclase activity partitioned with the photoreceptor outer segment (OS) cytoskeleton in a light-sensitive manner. RetGC-1 was found to bind actin filaments in actin blot overlays, suggesting a mechanism for its association with the OS cytoskeleton. In retinal sections, this enzyme was immunodetected only in the OSs, where it appeared to be distributed throughout the disk membranes.

scholarly journals CEP290 myosin-tail homology domain is essential for protein confinement between inner and outer segments in photoreceptors

2019 ◽  
Author(s):  
Poppy Datta ◽  
Brandon Hendrickson ◽  
Sarah Brendalen ◽  
Avri Ruffcorn ◽  
Seongjin Seo
Keyword(s):  
Membrane Proteins ◽  
Retinal Degeneration ◽  
Protein Trafficking ◽  
Outer Segment ◽  
Retinal Degenerations ◽  
Outer Segments ◽  
Disease Mechanisms ◽  
Initial Stage ◽  
Conditional Mutant ◽  
Ciliary Protein

ABSTRACTMutations in CEP290 cause various ciliopathies involving retinal degeneration. CEP290 proteins localize to the ciliary transition zone and are thought to act as a gatekeeper that controls ciliary protein trafficking. However, precise roles of CEP290 in photoreceptors and pathomechanisms of retinal degeneration in CEP290-associated ciliopathies are not sufficiently understood. Using Cep290 conditional mutant mice, in which the C-terminal myosin-tail homology domain is disrupted after the connecting cilium is assembled, we show that CEP290, more specifically the myosin-tail homology domain of CEP290, is essential for protein confinement between the inner and the outer segments. Inner segment plasma membrane proteins including STX3, SNAP25, and IMPG2 rapidly accumulate in the outer segment upon disruption of the myosin-tail homology domain. In contrast, localization of endomembrane proteins is not altered. Trafficking and confinement of most outer segment-resident proteins appear to be unaffected or only minimally affected in this mouse model. One notable exception is RHO, which exhibits severe mislocalization to inner segments from the initial stage of degeneration. Similar mislocalization phenotypes were observed in rd16 mice. These results suggest that failure of protein confinement at the connecting cilium and consequent accumulation of inner segment membrane proteins in the outer segment combined with insufficient RHO delivery is part of the disease mechanisms that cause retinal degeneration in CEP290-associated ciliopathies. Our study provides insights into the pathomechanisms of retinal degenerations associated with compromised ciliary gates.

scholarly journals Usher syndrome type 1–associated cadherins shape the photoreceptor outer segment

2017 ◽  
Vol 216 (6) ◽  
pp. 1849-1864 ◽  
Author(s):  
Cataldo Schietroma ◽  
Karine Parain ◽  
Amrit Estivalet ◽  
Asadollah Aghaie ◽  
Jacques Boutet de Monvel ◽  
...  
Keyword(s):  
Outer Segment ◽  
Usher Syndrome ◽  
Retinal Dystrophy ◽  
Syndrome Type ◽  
Photoreceptor Outer Segment ◽  
Outer Segments ◽  
Protocadherin 15 ◽  
Cadherin 23 ◽  
Usher Syndrome Type

Usher syndrome type 1 (USH1) causes combined hearing and sight defects, but how mutations in USH1 genes lead to retinal dystrophy in patients remains elusive. The USH1 protein complex is associated with calyceal processes, which are microvilli of unknown function surrounding the base of the photoreceptor outer segment. We show that in Xenopus tropicalis, these processes are connected to the outer-segment membrane by links composed of protocadherin-15 (USH1F protein). Protocadherin-15 deficiency, obtained by a knockdown approach, leads to impaired photoreceptor function and abnormally shaped photoreceptor outer segments. Rod basal outer disks displayed excessive outgrowth, and cone outer segments were curved, with lamellae of heterogeneous sizes, defects also observed upon knockdown of Cdh23, encoding cadherin-23 (USH1D protein). The calyceal processes were virtually absent in cones and displayed markedly reduced F-actin content in rods, suggesting that protocadherin-15–containing links are essential for their development and/or maintenance. We propose that calyceal processes, together with their associated links, control the sizing of rod disks and cone lamellae throughout their daily renewal.

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