Ablation of mpeg+ Macrophages Exacerbates mfrp-Related Hyperopia.

Purpose: Proper refractive development of the eye, termed emmetropization, is critical for focused vision and is impacted by both genetic determinants and several visual environment factors. Improper emmetropization caused by genetic variants can lead to congenital hyperopia, which is characterized by small eyes and relatively short ocular axial length. To date, variants in only four genes have been firmly associated with human hyperopia, one of which is MFRP.

Peripheral nerve pathology in sickle cell disease mice.

Introduction: Many patients with sickle cell disease (SCD) suffer from chronic pain, which is often described as neuropathic in nature. Although vascular and inflammatory pathology undoubtedly contribute to the SCD pain experience, the nociceptive signals that ultimately drive symptoms are detected and transmitted by peripheral sensory neurons. To date, no systematic histological examination of peripheral nerves has been completed in patients or mouse models of SCD to diagnose disease-related neuropathy.

PRCD is essential for high-fidelity photoreceptor disc formation.

Progressive rod-cone degeneration (PRCD) is a small protein residing in the light-sensitive disc membranes of the photoreceptor outer segment. Until now, the function of PRCD has remained enigmatic despite multiple demonstrations that its mutations cause blindness in humans and dogs. Here, we generated a PRCD knockout mouse and observed a striking defect in disc morphogenesis, whereby newly forming discs do not properly flatten.

Sustained Anti-Vascular Endothelial Growth Factor Activity of Aflibercept (Eylea) After Storage in Polycarbonate Syringes Used for Intravitreal Injection: A Pathway to Safety and Efficiency.

Aflibercept (Eylea™, Regeneron) is supplied in single-use glass vials along with 1 cc polycarbonate syringes. We sought to determine if storage of aflibercept for sustained periods within these syringes would result in loss of antivascular endothelial growth factor (anti-VEGF) activity. Aflibercept samples were drawn from commercially available glass vials into manufacturer-supplied 1-mL syringes and stored at 4°C.

Kif17 phosphorylation regulates photoreceptor outer segment turnover.

Background: KIF17, 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.

Cone myoid elongation involves unidirectional microtubule movement mediated by dynein-1.

Teleosts and amphibians exhibit retinomotor movements, morphological changes in photoreceptors regulated by light and circadian rhythms. Cone myoid elongation occurs during dark adaptation, leading to the positioning of the cone outer segment closer to the retinal pigment epithelium. Although it has been shown that microtubules are essential for cone myoid elongation, the underlying mechanism has not been established.

Loss of Zebrafish Mfrp Causes Nanophthalmia, Hyperopia, and Accumulation of Subretinal Macrophages.

Purpose: Mutations in membrane frizzled-related protein (MFRP) are associated with nanophthalmia, hyperopia, foveoschisis, irregular patches of RPE atrophy, and optic disc drusen in humans. Mouse mfrp mutants show retinal degeneration but no change in eye size or refractive state. The goal of this work was to generate zebrafish mutants to investigate the loss of Mfrp on eye size and refractive state, and to characterize other phenotypes observed.

The Retina and Other Light-sensitive Ocular Clocks.

Ocular clocks, first identified in the retina, are also found in the retinal pigment epithelium (RPE), cornea, and ciliary body. The retina is a complex tissue of many cell types and considerable effort has gone into determining which cell types exhibit clock properties. Current data suggest that photoreceptors as well as inner retinal neurons exhibit clock properties with photoreceptors dominating in nonmammalian vertebrates and inner retinal neurons dominating in mice.

Yap and Taz regulate retinal pigment epithelial cell fate.

The optic vesicle comprises a pool of bi-potential progenitor cells from which the retinal pigment epithelium (RPE) and neural retina fates segregate during ocular morphogenesis. Several transcription factors and signaling pathways have been shown to be important for RPE maintenance and differentiation, but an understanding of the initial fate specification and determination of this ocular cell type is lacking. We show that Yap/Taz-Tead activity is necessary and sufficient for optic vesicle progenitors to adopt RPE identity in zebrafish.

Subretinal infiltration of monocyte derived cells and complement misregulation in mice with AMD-like pathology.

We have characterized a naturally-occurring mutation in mice that causes slow, progressive photoreceptor degeneration, white fundus flecks, and late-onset RPE atrophy. These animals predictably lose visual function as photoreceptors degenerate. Genetic studies identified a deletion in the 5' coding sequence of Mfrp, designated Mfrp (174delG) , which essentially results in a complete knockout at the protein level.

The kinesin superfamily protein KIF17: one protein with many functions.

Kinesins are ATP-dependent molecular motors that carry cargos along microtubules, generally in an anterograde direction. They are classified into 14 distinct families with varying structural and functional characteristics. KIF17 is a member of the kinesin-2 family that is plus end-directed.

Kinesin-2 family motors in the unusual photoreceptor cilium.

This review focuses on recent advances in the understanding of kinesin-2 family motors in vertebrate photoreceptor development. Zebrafish photoreceptors develop by the 3rd day of embryogenesis, making it possible to study mutant phenotypes without the use of conditional alleles. Recent work using a zebrafish kif3b mutant allele validates the concept that the heterotrimeric kinesin II motor is generally required for ciliogenesis.

Analysis of KIF17 distal tip trafficking in zebrafish cone photoreceptors.

Multiple proteins are targeted to photoreceptor outer segments (OSs) where they function in phototransduction. Intraflagellar transport (IFT), a highly conserved bidirectional transport pathway occurring along the microtubules of the ciliary axoneme has been implicated in OS trafficking. The canonical anterograde motor for IFT is the heterotrimeric kinesin II or KIF3 complex.

The de-ubiquitinylating enzyme, USP2, is associated with the circadian clockwork and regulates its sensitivity to light.

We have identified a novel component of the circadian clock that regulates its sensitivity to light at the evening light to dark transition. USP2 (Ubiquitin Specific Protease 2), which de-ubiquitinylates and stabilizes target proteins, is rhythmically expressed in multiple tissues including the SCN. We have developed a knockout model of USP2 and found that exposure to low irradiance light at ZT12 increases phase delays of USP2(-/-) mice compared to wildtype.

174delG mutation in mouse MFRP causes photoreceptor degeneration and RPE atrophy.

Purpose: The authors have identified a recessive mutation causing progressive retinal degeneration, white fundus flecks, and eventual retinal pigment epithelium (RPE) atrophy. The goal of these studies was to characterize the retinal phenotype, to identify the causative locus, and to examine possible functions of the affected gene.

Methods: SNP mapping, DNA sequencing, and genetic complementation were used to identify the affected locus.

Functional and behavioral restoration of vision by gene therapy in the guanylate cyclase-1 (GC1) knockout mouse.

Background: Recessive mutations in guanylate cyclase-1 (Gucy2d) are associated with severe, early onset Leber congenital amaurosis-1(LCA1). Gucy2d encodes guanylate cyclase (GC1) is expressed in photoreceptor outer segment membranes and produces cGMP in these cells. LCA1 patients present in infancy with severely impaired vision and extinguished electroretinogram (ERG) but retain some photoreceptors in both their macular and peripheral retina for years.

Analysis of a zebrafish dync1h1 mutant reveals multiple functions for cytoplasmic dynein 1 during retinal photoreceptor development.

Background: Photoreceptors of the retina are highly compartmentalized cells that function as the primary sensory neurons for receiving and initiating transmission of visual information. Proper morphogenesis of photoreceptor neurons is essential for their normal function and survival. We have characterized a zebrafish mutation, cannonball, that completely disrupts photoreceptor morphogenesis.

Different roles for KIF17 and kinesin II in photoreceptor development and maintenance.

Kinesin 2 family members are involved in transport along ciliary microtubules. In Caenorhabditis elegans channel cilia, kinesin II and OSM-3 cooperate along microtubule doublets of the axoneme middle segment, whereas OSM-3 alone works on microtubule singlets to elongate the distal segment. Among sensory cilia, vertebrate photoreceptors share a similar axonemal structure with C.

Photoreceptor IFT complexes containing chaperones, guanylyl cyclase 1 and rhodopsin.

Intraflagellar transport (IFT) provides a mechanism for the transport of cilium-specific proteins, but the mechanisms for linkage of cargo and IFT proteins have not been identified. Using the sensory outer segments (OS) of photoreceptors, which are derived from sensory cilia, we have identified IFT-cargo complexes containing IFT proteins, kinesin 2 family proteins, two photoreceptor-specific membrane proteins, guanylyl cyclase 1 (GC1, Gucy2e) and rhodopsin (RHO), and the chaperones, mammalian relative of DNAJ, DnajB6 (MRJ), and HSC70 (Hspa8). Analysis of these complexes leads to a model in which MRJ through its binding to IFT88 and GC1 plays a critical role in formation or stabilization of the IFT-cargo complexes.

Analysis of IFT kinesins in developing zebrafish cone photoreceptor sensory cilia.

The photoreceptor outer segment (OS), a well-defined sensory cilium, provides an important context for the study of intraflagellar transport (IFT). The early phases of OS development involve successive events that are common to virtually all cilia. Additionally, intense protein trafficking occurs through the cilium and relies on IFT to maintain proper cellular morphology and optimize the photosensitive function.

Intraflagellar transport and the sensory outer segment of vertebrate photoreceptors.

Analysis of the other segments of rod and cone photoreceptors in vertebrates has provided a rich molecular understanding of how light absorbed by a visual pigment can result in changes in membrane polarity that regulate neurotransmitter release. These events are carried out by a large group of phototransduction proteins that are enriched in the outer segment. However, the mechanisms by which phototransduction proteins are sequestered in the outer segment are not well defined.

The homodimeric kinesin, Kif17, is essential for vertebrate photoreceptor sensory outer segment development.

Sensory cilia and intraflagellar transport (IFT), a pathway essential for ciliogenesis, play important roles in embryonic development and cell differentiation. In vertebrate photoreceptors IFT is required for the early development of ciliated sensory outer segments (OS), an elaborate organelle that sequesters the many proteins comprising the phototransduction machinery. As in other cilia and flagella, heterotrimeric members of the kinesin 2 family have been implicated as the anterograde IFT motor in OS.

Spatial distribution of intraflagellar transport proteins in vertebrate photoreceptors.

Intraflagellar transport (IFT) of a approximately 17S particle containing at least 16 distinct polypeptides is required for the assembly and maintenance of cilia and flagella. Although both genetic and biochemical evidence suggest a role for IFT in vertebrate photoreceptors, the spatial distribution of IFT proteins within photoreceptors remains poorly defined. We have evaluated the distribution of 4 IFT proteins using a combination of immunocytochemistry and rod-specific overexpression of GFP tagged IFT proteins.

Loss of Nocturnin, a circadian deadenylase, confers resistance to hepatic steatosis and diet-induced obesity.

The mammalian circadian system consists of a central oscillator in the suprachiasmatic nucleus of the hypothalamus, which coordinates peripheral clocks in organs throughout the body. Although circadian clocks control the rhythmic expression of a large number of genes involved in metabolism and other aspects of circadian physiology, the consequences of genetic disruption of circadian-controlled pathways remain poorly defined. Here we report that the targeted disruption of Nocturnin (Ccrn4l) in mice, a gene that encodes a circadian deadenylase, confers resistance to diet-induced obesity.