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 The outer segment serves as a default destination for the trafficking of membrane proteins in photoreceptors

2008 ◽  
Vol 183 (3) ◽  
pp. 485-498 ◽  
Author(s):  
Sheila A. Baker ◽  
Mohammad Haeri ◽  
Peter Yoo ◽  
Sidney M. Gospe ◽  
Nikolai P. Skiba ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Proteins ◽  
Outer Segment ◽  
The Other ◽  
Visual Signals ◽  
Its Sequence ◽  
Specific Targeting ◽  
New Material ◽  
Targeting Signals ◽  
Syntaxin 3

Photoreceptors are compartmentalized neurons in which all proteins responsible for evoking visual signals are confined to the outer segment. Yet, the mechanisms responsible for establishing and maintaining photoreceptor compartmentalization are poorly understood. Here we investigated the targeting of two related membrane proteins, R9AP and syntaxin 3, one residing within and the other excluded from the outer segment. Surprisingly, we have found that only syntaxin 3 has targeting information encoded in its sequence and its removal redirects this protein to the outer segment. Furthermore, proteins residing in the endoplasmic reticulum and mitochondria were similarly redirected to the outer segment after removing their targeting signals. This reveals a pattern where membrane proteins lacking specific targeting information are delivered to the outer segment, which is likely to reflect the enormous appetite of this organelle for new material necessitated by its constant renewal. This also implies that every protein residing outside the outer segment must have a means to avoid this “default” trafficking flow.

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 Accumulation of non-outer segment proteins in the outer segment underlies photoreceptor degeneration in Bardet–Biedl syndrome

2015 ◽  
Vol 112 (32) ◽  
pp. E4400-E4409 ◽  
Author(s):  
Poppy Datta ◽  
Chantal Allamargot ◽  
Joseph S. Hudson ◽  
Emily K. Andersen ◽  
Sajag Bhattarai ◽  
...  
Keyword(s):  
Protein Trafficking ◽  
Outer Segment ◽  
Primary Cilia ◽  
Leucine Zipper ◽  
Photoreceptor Cells ◽  
Photoreceptor Degeneration ◽  
Photoreceptor Outer Segment ◽  
Bardet Biedl Syndrome ◽  
Major Function ◽  
Underlying Mechanisms

Compartmentalization and polarized protein trafficking are essential for many cellular functions. The photoreceptor outer segment (OS) is a sensory compartment specialized for phototransduction, and it shares many features with primary cilia. As expected, mutations disrupting protein trafficking to cilia often disrupt protein trafficking to the OS and cause photoreceptor degeneration. Bardet–Biedl syndrome (BBS) is one of the ciliopathies associated with defective ciliary trafficking and photoreceptor degeneration. However, precise roles of BBS proteins in photoreceptor cells and the underlying mechanisms of photoreceptor degeneration in BBS are not well understood. Here, we show that accumulation of non-OS proteins in the OS underlies photoreceptor degeneration in BBS. Using a newly developed BBS mouse model [Leucine zipper transcription factor-like 1 (Lztfl1)/Bbs17 mutant], isolated OSs, and quantitative proteomics, we determined 138 proteins that are enriched more than threefold in BBS mutant OS. In contrast, only eight proteins showed a more than threefold reduction. We found striking accumulation of Stx3 and Stxbp1/Munc18-1 and loss of polarized localization of Prom1 within the Lztfl1 and Bbs1 mutant OS. Ultrastructural analysis revealed that large vesicles are formed in the BBS OS, disrupting the lamellar structure of the OS. Our findings suggest that accumulation (and consequent sequestration) of non-OS proteins in the OS is likely the primary cause of photoreceptor degeneration in BBS. Our data also suggest that a major function of BBS proteins in photoreceptors is to transport proteins from the OS to the cell body or to prevent entry of non-OS proteins into 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 Multistep peripherin-2/rds self-assembly drives membrane curvature for outer segment disk architecture and photoreceptor viability

2020 ◽  
Vol 117 (8) ◽  
pp. 4400-4410 ◽  
Author(s):  
Michelle L. Milstein ◽  
Breyanna L. Cavanaugh ◽  
Nicole M. Roussey ◽  
Stefanie Volland ◽  
David S. Williams ◽  
...  
Keyword(s):  
Self Assembly ◽  
Outer Segment ◽  
Structural Integrity ◽  
Normal Structure ◽  
Membrane Curvature ◽  
Structural Defects ◽  
Loss Of Function ◽  
Photoreceptor Outer Segment ◽  
Transgenic Expression ◽  
Curvature Continuity

Rod and cone photoreceptor outer segment (OS) structural integrity is essential for normal vision; disruptions contribute to a broad variety of retinal ciliopathies. OSs possess many hundreds of stacked membranous disks, which capture photons and scaffold the phototransduction cascade. Although the molecular basis of OS structure remains unresolved, recent studies suggest that the photoreceptor-specific tetraspanin, peripherin-2/rds (P/rds), may contribute to the highly curved rim domains at disk edges. Here, we demonstrate that tetrameric P/rds self-assembly is required for generating high-curvature membranes in cellulo, implicating the noncovalent tetramer as a minimal unit of function. P/rds activity was promoted by disulfide-mediated tetramer polymerization, which transformed localized regions of curvature into high-curvature tubules of extended lengths. Transmission electron microscopy visualization of P/rds purified from OS membranes revealed disulfide-linked tetramer chains up to 100 nm long, suggesting that chains maintain membrane curvature continuity over extended distances. We tested this idea in Xenopus laevis photoreceptors, and found that transgenic expression of nonchain-forming P/rds generated abundant high-curvature OS membranes, which were improperly but specifically organized as ectopic incisures and disk rims. These striking phenotypes demonstrate the importance of P/rds tetramer chain formation for the continuity of rim formation during disk morphogenesis. Overall, this study advances understanding of the normal structure and function of P/rds for OS architecture and biogenesis, and clarifies how pathogenic loss-of-function mutations in P/rds cause photoreceptor structural defects to trigger progressive retinal degenerations. It also introduces the possibility that other tetraspanins may generate or sense membrane curvature in support of diverse biological functions.

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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