(f) Presynaptic localization of mouse CaBP4 (red) demonstrated by covisualization with a presynaptic protein (PSD95; green). Arrows indicate rod spherules and cone pedicles. (g) Colocalization of synaptic vesicle proteins SV2 and synaptophysin (green) with mouse CaBP4 (red).
Biochemical evidence showing mouse CaBP4 (red) localization to the presynaptic region based on co-localization with the PSD95 marker (green).
RPGR was found in the connecting cilia of rods and cones with no evidence for species-dependent variation. RPGR ORF15 was the predominant variant in photoreceptor connecting cilia whereas constitutive RPGR (default) was the sole variant in the transitional zone of motile cilia in airway epithelia. Removal of soluble materials in the interphotoreceptor matrix facilitated detection of RPGR in the connecting cilia in photoreceptors. conclusions. RPGR localizes in photoreceptor connecting cilia and in a homologous structure, the transitional zone of motile cilia. These data are important for understanding the multitude of clinical manifestations associated with mutations in RPGR.
Selection B - Gene function is consistent with phenotype.
GOterm: GO:0032391 (photoreceptor connecting cilium)
FunctionalEvidence: This study uses a RPGR antibody to stain mouse retinal sections, showing localization of RPGR to the region between the outer segment and inner segment associated with the connecting cilia of rod and cone photoreceptors (Figure 2A). A similar experiment was performed in earlier studies with similar results (PMID: 11104772 Figure 4A, PMID: 10725384 Figure 2D upper). Within mouse airway epithelium tissue sections, RPGR antibody stained a narrow band across the apical domains of multi-ciliated cells (Figure 7A), corresponding with the basal parts of the motile cilia adjacent to the basal bodies (Figures 7B, 7C). This transitional zone of the motile cilia is the structural equivalent of the connecting cilia of photoreceptors, consistent with a biochemical role for RPGR in regulating ciliary organization and/or transport in both the retina and the respiratory tract.
FunctionalHPO: HP:0007642 (Congenital stationary night blindness), HP:0007994 (Peripheral visual field loss), HP:0000529 (Progressive visual loss), HP:0000365 (Hearing impairment), HP:0002205 (Recurrent respiratory infections), HP:0011109 (Chronic sinusitis), HP:0000388 (Otitis media), HP:0002837 (Recurrent bronchitis), HP:0012263 (Immotile cilia), HP:0005938 (Abnormal respiratory motile cilium morphology)
PhenotypeEvidence: This identification of RPGR as a potential regulator of ciliary organization and/or transport in multiple tissues is consistent with the diverse phenotypes associated with RPGR loss-of-function, ranging from visual defects associated with rod and cone dysfunction to respiratory defects associated with dysfunction of the respiratory motile cilia.
In cell-based assays, the overexpression of ADAM9 enhanced the ectodomain shedding of EphB4, Tie-2, Flk-1, CD40, VCAM, and VE-cadherin, so the enhanced expression of ADAM9 could potentially affect pathological neovascularization by increasing the shedding of these and other membrane proteins from endothelial cells. Finally, we provide the first evidence for the upregulation of ADAM9-dependent shedding by reactive oxygen species, which in turn are known to play a critical role in OIR.
Selection B- Gene function is consistent with phenotype.
Title: ADAM9 enhances protelytic shedding of membrane protein ectodomains.
GOterm: GO:0006509 (membrane protein ectodomain proteolysis), GO:0045765 (regulation of angiogenesis)
FunctionalEvidence: ADAM9 over-expression in COS-7 cells enhances proteolytic shedding of membrane proteins involved in angiogenesis (Figure 5). This effect is further enhanced by reactive oxygen species (Figures 6, 7).
FunctionalHPO: HP:0007843 (Attenuation of retinal blood vessels), HP:0000546 (Retinal degeneration)
PhenotypeEvidence: This evidence is consistent with attenuation of retinal blood vessels observed in human patients as well as the finding from this same paper that Adam9 knockout mice exhibit reduced ocular neovascularization (Figures 1, 3).
The interphotoreceptor matrix (IPM) occupies the extracellular space between the apical surface of the retinal pigmented epithelium and the external limiting membrane of the neural retina. This space contains two chondroitin sulfate proteoglycans, designated IPM 150 and IPM 200
Selection B- Gene function is consistent with phenotype.
GOterm: GO:0033165 (interphotoreceptor matrix)
FunctionalEvidence: IMPG1 protein can be purified as a component of an insoluble extracellular layer that is located between apical side of the RPE and the external limiting membrane of the neural retina, known as the interphotoreceptor matrix (Figure 1). This layer is known to support the function and maintenance of photoreceptor cells (PMID: 7344830).
FunctionalHPO: HP:0030612 (Abnormal retinal morphology on macular OCT, specifically photoreceptor outer segment elongation), HP:0007677 (Vitelliform-like macular lesions)
PhenotypeEvidence: The article provides an example of the function of IMPG1 (here referred to as IPM150) as a component of the interphotoreceptor matrix, an insoluble extracellular layer that is located between apical side of the RPE and the external limiting membrane of the neural retina, and which supports the adhesion, function, and maintenance of photoreceptor cells.
The disintegrin domain of MDC-9 lacks an RGD motif but has recently been reported to bind the alpha(6)beta(1) integrin; however, it is unclear whether MDC-9 can bind other integrins. In the present study myeloma cells, but not lymphoblastoid cells, were shown to bind to immobilised, recombinantly expressed MDC-9 disintegrin domain (A9dis). Binding was divalent cation-dependent, being supported by Mn(2+) and Ca(2+). Adhesion of myeloma cells to A9dis was completely inhibited by an antibody to the alpha(v)beta(5) integrin but not by antibodies to other subunits. RGD-containing peptides had no effect on binding, suggesting that MDC-9 interacts with alpha(v)beta(5) in an RGD-independent manner. Flow cytometric analyses demonstrated that myeloma cells, but not lymphoblastoid cells, expressed alpha(v)beta(5) on the cell membrane. These data indicated that the disintegrin domain of MDC-9 can function as an adhesion molecule by interacting with an alpha(v)beta(5) integrin.
Selection B- Gene function is consistent with phenotype.
Title: ADAM9 (MDC-9) functions as an adhesion molecule by interacting with integrins.
GOterm: GO:0033630 (positive regulation of cell adhesion mediated by integrin)
FunctionalEvidence: Melanoma cells bind to they the immobilized recombinant ADAM9 disintegrin domain in a manner that is dependent on divalent cations (Mn2+ or Ca2+). The interaction can also be disrupted by an antibody to the alpha(v)beta(5) integrin, indicating that the cell-cell adhesion function of ADAM9 is mediated by an interaction with this molecule.
FunctionalHPO: HP:0000546 (Retinal degeneration), HP:0007703 (Abnormality of retinal pigmentation), HP:0030610 (Photoreceptor outer segment loss on macular OCT
PhenotypeEvidence: This evidence is consistent with evidence that Adam9 knockout mice exhibit a gap between the photoreceptor outer nuclear segment and the retinal pigment epithelium, indicating that loss of this adhesion molecule may impair the integrity of retinal organization.
We provide here structural evidence that the protein product of the ocular albinism type 1 gene (OA1), a pigment cell-specific integral membrane glycoprotein, represents a novel member of the GPCR superfamily and demonstrate that it binds heterotrimeric G proteins. Moreover, we show that OA1 is not found at the plasma membrane, being instead targeted to specialized intracellular organelles, the melanosomes. Our data suggest that OA1 represents the first example of an exclusively intracellular GPCR
Selection B- Gene function is consistent with phenotype.
Title: GPR143 binds heterotrimeric G proteins and shows subcellular localization to melanosomes.
GOterm: GO:0032402 (melanosome transport)
FunctionalEvidence: Exogenously expressed GPR143 in COS-7 cells forms protein-protein interactions with heterotrimeric G-proteins that can be co-immunoprecipitated (Figure 2). This was validated with endogenous GPR143 (Figure 3). GPR143 shows subcellular localization not to the plasma membrane but to melanosomes (Figures 4, 6).
FunctionalHPO: HP:0005592 (Giant melanosomes in melanocytes), severely reduced microtubule-based motility in melanosomes
HumanPhenotype: HP:0005592 (Giant melanosomes in melanocytes), HP:0007894 (Hypopigmentation of the fundus), severely reduced microtubule-based motility in melanosomes, Giant melanosomes in retinal pigment epithelial cells
PhenotypeEvidence: This evidence is consistent with the melanocyte trafficking defects characteristic of human patients.
Protein extracts from wild-type and LAP-BBS4 transgenic testes were subjected to tandem affinity purification. In LAP-BBS4 transgenic testis, all BBSome subunits (Bbs1, Bbs2, Bbs5, Bbs7, Bbs8, and Bbs9) were co-purified with LAP-BBS4, while no BBS proteins were purified when wild-type testes were used (Figure 1A and Table S1).
Selection B- Gene function is consistent with phenotype.
GOterm: GO:0034464 (BBSome)
FunctionalEvidence: Affinity-purified human BBS4 expressed from a transgene in mouse testes resulted in co-purification of all BBSome subunits, including Bbs1, as identified by mass spectrometry (Figure 1A).
FunctionalHPO: HP:0000026 (Male hypogonadism), HP:0000510 (Rod-cone dystrophy), HP:0001513 (Obesity)
Subcellular localization of spam to the outer segment of mature porcine retina using antibody to spam (green) and antibody to rhodopsin (red). The overlay shows the localization of spam in the same region as rhodopsin (yellow) in the outer segment of the photoreceptor cell layer.
Selection B- Gene function is consistent with phenotype.
GOterm: GO:0001750 (cellular component of the photoreceptor outer segment)
FunctionalEvidence: Immunofluorescent staining of mature porcine retina tissues with an antibody to the EYS protein shows subcellular localization to the outer segment specifically (Figure 2b). This region of the retina holds the light-absorbing, ciliated part of the photoreceptor cells, and localization / function in this region is consistent with the rod and cone photoreceptor dysfunction phenotypes found in the human patients.
FunctionalHPO: Nyctalopia HP:0000662, Undetectable electroretinogram HP:0000550, Reduced visual acuity HP:0007663
Glycosaminoglycans: KS and C6S
ExperimentName: Accumulation of glycosaminoglycans
GOterm:
FunctionalEvidence: Patient cells assayed with artificial substrates for the N-acetylgalactosamine-6-sulfate sulfatase encoded by GALNS, depending on the severity of the coding sequence alteration, are either unable to break down the substrate or are hindered. Further, in mucopolysaccharidosis IVA patients with known variants, elevated levels of keratan sulfate and chondroitin-6-sulfate can be found in their urine where levels in unaffected individuals are much lower.
HGNC:
GeneEvidence:
SharedGeneImplication:
Glycosaminoglycans: KS and C6S
ExperimentName: Accumulation of glycosaminoglycans
GOterm:
FunctionalEvidence: Patient cells assayed with artificial substrates for the N-acetylgalactosamine-6-sulfate sulfatase encoded by GALNS, depending on the severity of the coding sequence alteration, are either unable to break down the substrate or are hindered. Further, in mucopolysaccharidosis IVA patients with known variants, elevated levels of keratan sulfate and chondroitin-6-sulfate can be found in their urine where levels in unaffected individuals are much lower.
HGNC:
GeneEvidence:
SharedGeneImplication: