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.

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 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 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 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.

scholarly journals Syntaxin 3 is essential for photoreceptor outer segment protein trafficking and survival

2020 ◽  
Vol 117 (34) ◽  
pp. 20615-20624
Author(s):  
Mashal Kakakhel ◽  
Lars Tebbe ◽  
Mustafa S. Makia ◽  
Shannon M. Conley ◽  
David M. Sherry ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Protein Trafficking ◽  
Outer Segment ◽  
Functional Decline ◽  
Structural Defects ◽  
Photoreceptor Outer Segment ◽  
Photoreceptor Membrane ◽  
Synaptic Region ◽  
Syntaxin 3

Trafficking of photoreceptor membrane proteins from their site of synthesis in the inner segment (IS) to the outer segment (OS) is critical for photoreceptor function and vision. Here we evaluate the role of syntaxin 3 (STX3), in trafficking of OS membrane proteins such as peripherin 2 (PRPH2) and rhodopsin. Photoreceptor-specificStx3knockouts [Stx3f/f(iCre75)andStx3f/f(CRX-Cre)] exhibited rapid, early-onset photoreceptor degeneration and functional decline characterized by structural defects in IS, OS, and synaptic terminals. Critically, in the absence of STX3, OS proteins such as PRPH2, the PRPH2 binding partner, rod outer segment membrane protein 1 (ROM1), and rhodopsin were mislocalized along the microtubules to the IS, cell body, and synaptic region. We find that the PRPH2 C-terminal domain interacts with STX3 as well as other photoreceptor SNAREs, and our findings indicate that STX3 is an essential part of the trafficking pathway for both disc (rhodopsin) and rim (PRPH2/ROM1) components of the OS.

scholarly journals Differential requirement of NPHP1 for compartmentalized protein localization during photoreceptor outer segment development and maintenance

2021 ◽  
Author(s):  
Poppy Datta ◽  
J. Thomas Cribbs ◽  
Seongjin Seo
Keyword(s):  
Plasma Membrane ◽  
Membrane Proteins ◽  
Retinal Degeneration ◽  
Outer Segment ◽  
Protein Localization ◽  
Retinal Dystrophy ◽  
Gene Trap ◽  
Mutant Mice ◽  
Plasma Membrane Proteins ◽  
Photoreceptor Outer Segment

AbstractNephrocystin (NPHP1) is a ciliary transition zone protein and its ablation causes nephronophthisis (NPHP) with partially penetrant retinal dystrophy. However, the precise requirements of NPHP1 in photoreceptors are not well understood. Here, we characterize retinal degeneration in a mouse model of NPHP1 and show that NPHP1 is required to prevent infiltration of inner segment plasma membrane proteins into the outer segment during the photoreceptor maturation. We demonstrate that Nphp1 gene-trap mutant mice, which were previously described as null, are in fact hypomorphs due to the production of a small quantity of functional mRNAs derived from nonsense-associated altered splicing and skipping of two exons including the one harboring the gene-trap. In homozygous mutant animals, inner segment plasma membrane proteins such as syntaxin-3 (STX3), synaptosomal-associated protein 25 (SNAP25), and interphotoreceptor matrix proteoglycan 2 (IMPG2) accumulate in the outer segment when outer segments are actively elongating. This phenotype, however, is spontaneously ameliorated after the outer segment elongation is completed. Retinal degeneration also occurs temporarily during the photoreceptor maturation but stops afterward. We further show that Nphp1 genetically interacts with Cep290, another NPHP gene, and that a reduction of Cep290 gene dose results in retinal degeneration that continues until adulthood in Nphp1 mutant mice. These findings demonstrate that NPHP1 is required for the confinement of inner segment plasma membrane proteins during the outer segment development, but its requirement diminishes as photoreceptors mature. Our study also suggests that additional mutations in other NPHP genes may influence the penetrance of retinopathy in human NPHP1 patients.

scholarly journals Ift172 conditional knockout mice exhibit rapid retinal degeneration and protein trafficking defects

2017 ◽  
Author(s):  
Priya R Gupta ◽  
Nachiket Pendse ◽  
Scott H Greenwald ◽  
Mihoko Leon ◽  
Qin Liu ◽  
...  
Keyword(s):  
Retinal Degeneration ◽  
Protein Trafficking ◽  
Primary Cilia ◽  
Molecular Mechanisms ◽  
Outer Nuclear Layer ◽  
Intraflagellar Transport ◽  
Conditional Knockout ◽  
Photoreceptor Outer Segments ◽  
Outer Segments ◽  
Trafficking Defects

AbstractIntraflagellar transport (IFT) is a bidirectional transport process that occurs along primary cilia and specialized sensory cilia, such as photoreceptor outer-segments. Genes coding for various IFT components are associated with ciliopathies. Mutations in IFT172 lead to diseases ranging from isolated retinal degeneration to severe syndromic ciliopathies. In this study, we created a mouse model of IFT172-associated retinal degeneration to investigate the ocular disease mechanism. We found that depletion of IFT172 in rod photoreceptors leads to a rapid degeneration of the retina, with severely reduced electroretinography responses by one month and complete outer-nuclear layer degeneration by two months. We investigated molecular mechanisms of degeneration and show that IFT172 protein reduction leads to mislocalization of specific photoreceptor outer-segments proteins (RHO, RP1, IFT139), aberrant light-driven translocation of alpha transducin and altered localization of glioma-associated oncogene family member 1 (GLI1). This murine model recapitulates the retinal phenotype seen in patients with IFT172-associated blindness and can be used for in vivo testing of ciliopathy therapies.

scholarly journals Rpgrip1 is required for rod outer segment development and ciliary protein trafficking in zebrafish

2017 ◽  
Vol 7 (1) ◽  
Author(s):  
Rakesh K. Raghupathy ◽  
Xun Zhang ◽  
Fei Liu ◽  
Reem H. Alhasani ◽  
Lincoln Biswas ◽  
...  
Keyword(s):  
Protein Trafficking ◽  
Outer Segment ◽  
Rod Outer Segment ◽  
Ciliary Protein

scholarly journals Tmem138, a photoreceptor connecting cilium (CC) protein, is required for rhodopsin transport across the cilium and outer segment (OS) biogenesis

2021 ◽  
Author(s):  
Dianlei Liu ◽  
Jiali Ru ◽  
Lijing Xie ◽  
Mingjuan Wu ◽  
Yingchun Su ◽  
...  
Keyword(s):  
Protein Trafficking ◽  
Outer Segment ◽  
Primary Cilia ◽  
Early Stage ◽  
Joubert Syndrome ◽  
Gene Encoding ◽  
Retinal Degenerations ◽  
Retinal Dystrophies ◽  
Broad Array ◽  
Transport Complex

Photoreceptor connecting cilium (CC) is structurally analogous to the transition zone (TZ) of primary cilia and gates the molecular trafficking between the inner and the outer segment (OS). Retinal dystrophies with underlying CC defects are manifested in a broad array of syndromic conditions known as ciliopathies as well as non-syndromic retinal degenerations. Despite extensive studies, protein trafficking across the photoreceptor CC is largely unknown. Here we genetically inactivated mouse Tmem138, a gene encoding a ciliary membrane protein localized to the ciliary TZ and linked to Joubert syndrome (JBTS). Germline deletion of Tmem138 abolished OS morphogenesis followed by rapid photoreceptor degeneration. Tmem138 was found localized to the photoreceptor CC and, accordingly, the molecular compartments of the CC and axoneme of the mutant photoreceptors were altered despite ciliogenesis proceeding normally at the early stage of photoreceptor development. To gain further insights into Tmem138 function in OS biogenesis, we focused on trafficking of rhodopsin, the most abundant protein of the OS. Mislocalization of rhodopsin was readily observed as early as P5 in the mutant photoreceptors prior to growth of the OS. Ablation of Tmem138 in mature rods recapitulated the molecular changes in the germline mutants, causing well-formed outer segment discs to disintegrate accompanied by mislocalization of rhodopsin in the cell body. Furthermore, Tmem138 interacted with rhodopsin, and two additional CC compartment proteins Ahi1 and Tmem231, which were both altered in the mutant photoreceptors. Taken together, these results suggest that Tmem138 has a distinct role in gating the transport of rhodopsin and likely other OS bound proteins through formation of CC transport complex(es).

scholarly journals Differential requirement of NPHP1 for compartmentalized protein localization during photoreceptor outer segment development and maintenance

PLoS ONE ◽  
2021 ◽  
Vol 16 (5) ◽  
pp. e0246358
Author(s):  
Poppy Datta ◽  
J. Thomas Cribbs ◽  
Seongjin Seo
Keyword(s):  
Plasma Membrane ◽  
Membrane Proteins ◽  
Retinal Degeneration ◽  
Outer Segment ◽  
Protein Localization ◽  
Retinal Dystrophy ◽  
Gene Trap ◽  
Mutant Mice ◽  
Plasma Membrane Proteins ◽  
Photoreceptor Outer Segment

Nephrocystin (NPHP1) is a ciliary transition zone protein and its ablation causes nephronophthisis (NPHP) with partially penetrant retinal dystrophy. However, the precise requirements of NPHP1 in photoreceptors are not well understood. Here, we characterize retinal degeneration in a mouse model of NPHP1 and show that NPHP1 is required to prevent infiltration of inner segment plasma membrane proteins into the outer segment during the photoreceptor maturation. We demonstrate that Nphp1 gene-trap mutant mice, which were previously described as null, are likely hypomorphs due to the production of a small quantity of functional mRNAs derived from nonsense-associated altered splicing and skipping of two exons including the one harboring the gene-trap. In homozygous mutant animals, inner segment plasma membrane proteins such as syntaxin-3 (STX3), synaptosomal-associated protein 25 (SNAP25), and interphotoreceptor matrix proteoglycan 2 (IMPG2) accumulate in the outer segment when outer segments are actively elongating. This phenotype, however, is spontaneously ameliorated after the outer segment elongation is completed. Consistent with this, some photoreceptor cell loss (~30%) occurs during the photoreceptor maturation period but it stops afterward. We further show that Nphp1 genetically interacts with Cep290, another NPHP gene, and that a reduction of Cep290 gene dose results in retinal degeneration that continues until adulthood in Nphp1 mutant mice. These findings demonstrate that NPHP1 is required for the confinement of inner segment plasma membrane proteins during the outer segment development, but its requirement diminishes as photoreceptors mature. Our study also suggests that additional mutations in other NPHP genes may influence the penetrance of retinopathy in human NPHP1 patients.

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