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.

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 AAV Vectors for FRET-Based Analysis of Protein-Protein Interactions in Photoreceptor Outer Segments

2016 ◽  
Vol 10 ◽  
Author(s):  
Elvir Becirovic ◽  
Sybille Böhm ◽  
Ong N. P. Nguyen ◽  
Lisa M. Riedmayr ◽  
Verena Hammelmann ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Protein Protein Interactions ◽  
Photoreceptor Outer Segments ◽  
Outer Segments

scholarly journals The Transcriptional Coactivator Cbp Interacts with β-Catenin to Activate Gene Expression

2000 ◽  
Vol 149 (2) ◽  
pp. 249-254 ◽  
Author(s):  
Ken-Ichi Takemaru ◽  
Randall T. Moon
Keyword(s):  
Protein Interactions ◽  
Transcriptional Activation ◽  
Target Genes ◽  
Terminal Region ◽  
Creb Binding Protein ◽  
Transcriptional Coactivator ◽  
Protein Protein Interactions ◽  
Yeast Two Hybrid System ◽  
Recruitment System

β-Catenin plays a pivotal role in the transcriptional activation of Wnt-responsive genes by binding to TCF/LEF transcription factors. Although it has been suggested that the COOH-terminal region of β-catenin functions as an activation domain, the mechanisms of activation remain unclear. To screen for potential transcriptional coactivators that bind to the COOH-terminal region of β-catenin, we used a novel yeast two-hybrid system, the Ras recruitment system (RRS) that detects protein–protein interactions at the inner surface of the plasma membrane. Using this system, we isolated the CREB-binding protein (CBP). Armadillo (Arm) repeat 10 to the COOH terminus of β-catenin is involved in binding to CBP, whereas β-catenin interacts directly with the CREB-binding domain of CBP. β-Catenin synergizes with CBP to stimulate the activity of a synthetic reporter in vivo. Conversely, β-catenin–dependent transcriptional activation is repressed by E1A, an antagonist of CBP function, but not by an E1A mutant that does not bind to CBP. The activation of Wnt target genes such as siamois and Xnr3 in Xenopus embryos is also sensitive to E1A. These findings suggest that CBP provides a link between β-catenin and the transcriptional machinery, and possibly mediates the oncogenic function of β-catenin.

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 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.

IL-14 MAPPIT: a versatile METHOD to study protein-protein interactions in mammalian cells

2003 ◽  
Vol 16 (5) ◽  
pp. 577-577
Author(s):  
J. Tavernier ◽  
S. Eyckerman ◽  
I. Lemmens ◽  
S. Lievens ◽  
J. Vandekerckhove ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Mammalian Cells ◽  
Protein Protein Interactions

scholarly journals Distinct p63 and p73 Protein Interactions Predict Specific Functions in mRNA Splicing and Polyploidy Control in Epithelia

Cells ◽  
2020 ◽  
Vol 10 (1) ◽  
pp. 25
Author(s):  
Julian M. Rozenberg ◽  
Olga S. Rogovaya ◽  
Gerry Melino ◽  
Nickolai A. Barlev ◽  
Alexander Kagansky
Keyword(s):  
Protein Interactions ◽  
Transcriptional Repression ◽  
Genome Stability ◽  
Spindle Assembly Checkpoint ◽  
Genotoxic Stress ◽  
Protein Protein Interactions ◽  
Proliferating Cells ◽  
P53 Family ◽  
Differentiated Cells ◽  
Specific Localization

Epithelial organs are the first barrier against microorganisms and genotoxic stress, in which the p53 family members p63 and p73 have both overlapping and distinct functions. Intriguingly, p73 displays a very specific localization to basal epithelial cells in human tissues, while p63 is expressed in both basal and differentiated cells. Here, we analyse systematically the literature describing p63 and p73 protein–protein interactions to reveal distinct functions underlying the aforementioned distribution. We have found that p73 and p63 cooperate in the genome stability surveillance in proliferating cells; p73 specific interactors contribute to the transcriptional repression, anaphase promoting complex and spindle assembly checkpoint, whereas p63 specific interactors play roles in the regulation of mRNA processing and splicing in both proliferating and differentiated cells. Our analysis reveals the diversification of the RNA and DNA specific functions within the p53 family.

scholarly journals β-Catenin is a pH sensor with decreased stability at higher intracellular pH

2018 ◽  
Vol 217 (11) ◽  
pp. 3965-3976 ◽  
Author(s):  
Katharine A. White ◽  
Bree K. Grillo-Hill ◽  
Mario Esquivel ◽  
Jobelle Peralta ◽  
Vivian N. Bui ◽  
...  
Keyword(s):  
Intracellular Ph ◽  
Protein Interactions ◽  
Mammalian Cells ◽  
Ph Sensor ◽  
Adherens Junction ◽  
Protein Protein Interactions ◽  
Adherens Junction Protein ◽  
Junction Protein ◽  
Evolutionarily Conserved ◽  
Ph Dependent

β-Catenin functions as an adherens junction protein for cell–cell adhesion and as a signaling protein. β-catenin function is dependent on its stability, which is regulated by protein–protein interactions that stabilize β-catenin or target it for proteasome-mediated degradation. In this study, we show that β-catenin stability is regulated by intracellular pH (pHi) dynamics, with decreased stability at higher pHi in both mammalian cells and Drosophila melanogaster. β-Catenin degradation requires phosphorylation of N-terminal residues for recognition by the E3 ligase β-TrCP. While β-catenin phosphorylation was pH independent, higher pHi induced increased β-TrCP binding and decreased β-catenin stability. An evolutionarily conserved histidine in β-catenin (found in the β-TrCP DSGIHS destruction motif) is required for pH-dependent binding to β-TrCP. Expressing a cancer-associated H36R–β-catenin mutant in the Drosophila eye was sufficient to induce Wnt signaling and produced pronounced tumors not seen with other oncogenic β-catenin alleles. We identify pHi dynamics as a previously unrecognized regulator of β-catenin stability, functioning in coincidence with phosphorylation.

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