Photoreceptor damage following exposure to excess riboflavin

Flavins generate oxidants during metabolism and when exposed to light. Here we report that the photoreceptor layer of retinas from black-eyed rats is reduced in size by a dietary regime containing excess riboflavin. The effect of excess riboflavin was dose-dependent and was manifested by a decrease in photoreceptor length. This decrease was due in part to a reduction in the thickness of the outer nuclear layer, a structure formed from stacked photoreceptor nuclei. These changes were accompanied by an increase in photoreceptor outer segment autofluorescence following illumination at 328 nm, a wavelength that corresponds to the excitation maxima of oxidized lipopigments of the retinal pigment epithelium.     

Metabolic and redox signaling in the retina

Visual perception by photoreceptors relies on the interaction of incident photons from light with a derivative of vitamin A that is covalently linked to an opsin molecule located in a special subcellular structure, the photoreceptor outer segment. The photochemical reaction produced by the photon is optimal when the opsin molecule, a seven-transmembrane protein, is embedded in a lipid bilayer of optimal fluidity. This is achieved in vertebrate photoreceptors by a high proportion of lipids made with polyunsaturated fatty acids, which have the detrimental property of being oxidized and damaged by light. Photoreceptors cannot divide, but regenerate their outer segments. This is an enormous energetic challenge that explains why photoreceptors metabolize glucose through aerobic glycolysis, as cancer cells do. Uptaken glucose produces metabolites to renew that outer segment as well as reducing power through the pentose phosphate pathway to protect photoreceptors against oxidative damage.     

The absence of melanopsin alters retinal clock function and dopamine regulation by light

The retinal circadian clock is crucial for optimal regulation of retinal physiology and function, yet its cellular location in mammals is still controversial. We used laser microdissection to investigate the circadian profiles and phase relations of clock gene expression and Period gene induction by light in the isolated outer (rods/cones) and inner (inner nuclear and ganglion cell layers) regions in wild-type and melanopsin-knockout (Opn ₄ ^?/? ) mouse retinas. In the wild-type mouse, all clock genes are rhythmically expressed in the photoreceptor layer but not in the inner retina. For clock genes that are rhythmic in both retinal compartments, the circadian profiles are out of phase. These results are consistent with the view that photoreceptors are a potential site of circadian rhythm generation. In mice lacking melanopsin, we found an unexpected loss of clock gene rhythms and of the photic induction of Per1-Per2 mRNAs only in the outer retina. Since melanopsin ganglion cells are known to provide a feed-back signalling pathway for photic information to dopaminergic cells, we further examined dopamine (DA) synthesis in Opn ₄ ^?/? mice. The lack of melanopsin prevented the light-dependent increase of tyrosine hydroxylase (TH) mRNA and of DA and, in constant darkness, led to comparatively high levels of both components. These results suggest that melanopsin is required for molecular clock function and DA regulation in the retina, and that Period gene induction by light is mediated by a melanopsin-dependent, DA-driven signal acting on retinal photoreceptors.     

Soluble factors and the development of rod photoreceptors

Photoreceptors are the most abundant cell type in the vertebrate neural retina. Like the other retinal neurons and the Müller glia, they arise from a population of precursor cells that are multipotent and intrinsic to the retina. Approximately 10 years ago, several studies demonstrated that retinal precursor cells (RPCs) are competent to respond to environmental factors that promote cell type determination and differentiation. Since those studies, significant effort has been directed at identifying the molecular nature of these environmental signals and understanding the precise mechanisms they employ to drive RPCs towards the different retinal fates. In this review, we describe the recent progress toward understanding how environmental factors influence the development of vertebrate rod photoreceptors.     

Reprogramming the metabolome rescues retinal degeneration

Metabolomics studies in the context of ophthalmology have largely focused on identifying metabolite concentrations that characterize specific retinal diseases. Studies involving mass spectrometry (MS) and nuclear magnetic resonance (NMR) spectroscopy have shown that individuals suffering from retinal diseases exhibit metabolic profiles that markedly differ from those of control individuals, supporting the notion that metabolites may serve as easily identifiable biomarkers for specific conditions. An emerging branch of metabolomics resulting from biomarker studies, however, involves the study of retinal metabolic dysfunction as causes of degeneration. Recent publications have identified a number of metabolic processes—including but not limited to glucose and oxygen metabolism—that, when perturbed, play a role in the degeneration of photoreceptor cells. As a result, such studies have led to further research elucidating methods for prolonging photoreceptor survival in an effort to halt degeneration in its early stages. This review will explore the ways in which metabolomics has deepened our understanding of the causes of retinal degeneration and discuss how metabolomics can be used to prevent retinal degeneration from progressing to its later disease stages.     

Ultrastructure and biochemistry of the pineal organ in deep-sea lanternfishes (Myctophidae)

Pineal structural and biochemical adaptations in lanternfishes included: 1) few photoreceptor outer segment discs; 2) conventional synapses between photoreceptors and pineal neurons; and 3) low levels (0-60 pg/pineal) of serotonin compared to those (greater than 1.0 ng/pineal) in the goldfish pineal organ. These findings suggest reduced photosensory and/or neuroendocrine functions in these deep-sea fishes.     

Ultrastructure and biochemistry of the pineal organ in deep-sea lanternfishes (Myctophidae)

Summary Pineal structural and biochemical adaptations in lanternfishes included: 1) few photoreceptor outer segment discs; 2) conventional synapses between photoreceptors and pineal neurons; and 3) low levels (0–60 pg/pineal) of serotonin compared to those (>1.0 ng/pineal) in the goldfish pineal organ. These findings suggest reduced photosensory and/or neuroendocrine functions in these deep-sea fishes.     

Nesprin interchain associations control nuclear size

Nesprins-1/-2/-3/-4 are nuclear envelope proteins, which connect nuclei to the cytoskeleton. The largest nesprin-1/-2 isoforms (termed giant) tether F-actin through their N-terminal actin binding domain (ABD). Nesprin-3, however, lacks an ABD and associates instead to plectin, which binds intermediate filaments. Nesprins are integrated into the outer nuclear membrane via their C-terminal KASH-domain. Here, we show that nesprin-1/-2 ABDs physically and functionally interact with nesprin-3. Thus, both ends of nesprin-1/-2 giant are integrated at the nuclear surface: via the C-terminal KASH-domain and the N-terminal ABD-nesprin-3 association. Interestingly, nesprin-2 ABD or KASH-domain overexpression leads to increased nuclear areas. Conversely, nesprin-2 mini (contains the ABD and KASH-domain but lacks the massive nesprin-2 giant rod segment) expression yields smaller nuclei. Nuclear shrinkage is further enhanced upon nesprin-3 co-expression or microfilament depolymerization. Our findings suggest that multivariate intermolecular nesprin interactions with the cytoskeleton form a lattice-like filamentous network covering the outer nuclear membrane, which determines nuclear size.     

Signaling pathways and the cerebral cavernous malformations proteins: lessons from structural biology

Cerebral cavernous malformations (CCM) are neurovascular dysplasias that result in mulberry-shaped lesions predominantly located in brain and spinal tissues. Mutations in three genes are associated with CCM. These genes encode for the proteins KRIT1/CCM1 (krev interaction trapped 1/cerebral cavernous malformations 1), cerebral cavernous malformations 2, osmosensing scaffold for MEKK3 (CCM2/malcavernin/OSM), and cerebral cavernous malformations 3/programmed cell death 10 (CCM3/PDCD10). There have been many significant recent advances in our understanding of the structure and function of these proteins, as well as in their roles in cellular signaling. Here, we provide an update on the current knowledge of the structure of the CCM proteins and their functions within cellular signaling, particularly in cellular adhesion complexes and signaling cascades. We go on to discuss subcellular localization of the CCM proteins, the formation and regulation of the CCM complex signaling platform, and current progress towards targeted therapy for CCM disease. Recent structural studies have begun to shed new light on CCM protein function, and we focus here on how these studies have helped inform the current understanding of these roles and how they may aid future studies into both CCM-related biology and disease mechanisms.     

Further studies on the effect of red and far red light on rat retinal development

Summary The influence on the development of the outer segments of the rat retina of far red and red light in different sequences was investigated. The far red treatment appeared to be dominating, and for animals treated with far red light first, further treatment of red light could not bring the outer segment growth back to normal. The treatments also initiated different dopamine uptakes in the retinas.     

Enteropathogenic Escherichia coli: a pathogen that inserts its own receptor into host cells

Enteropathogenic Escherichia coli (EPEC) is a major cause of infant diarrhea, killing hundreds of thousands of children per year worldwide. Intimate attachment to the host cell leading to the formation of actin-rich pedestals beneath the adhering bacteria is an essential feature of EPEC pathogenesis. EPEC attaches to host cells via the outer membrane adhesin, intimin. It was recently shown that EPEC inserts its own receptor for intimate adherence, Tir (translocated intimin receptor) into the host cell membrane. The focus of this review is on the discovery and characterization of this novel receptor, and our current understanding of its role in pedestal formation. Gram-negative bacterial secretion systems, including type III secretion systems, are reviewed and discussed in the context of Tir delivery into the host cell membrane. The relationship and relevance of in vitro models compared to the actual in vivo situation is essential to understanding disease. We have critically reviewed the use of animal models in studying EPEC infection. Elucidating the function of Tir will contribute to our understanding of how EPEC mediates disease.     

The use of iodinated density gradient media for the isolation of rod outer segments

Sucrose, nycodenz, metrizamide and a mixture of equal volumes of sucrose and metrizamide were used as density gradient media for the isolation of retinal rod outer segments. The high osmolarity of sucrose had a strongly negative effect on the nature of the rod outer segments, whereas they were much better preserved using iodinated density gradient media such as nycodenz and metrizamide for their isolation.     

The use of iodinated density gradient media for the isolation of rod outer segments

Summary Sucrose, nycodenz, metrizamide and a mixture of equal volumes of sucrose and metrizamide were used as density gradient media for the isolation of retinal rod outer segments. The high osmolarity of sucrose had a strongly negative effect on the nature of the rod outer segments, whereas they were much better preserved using iodinated density gradient media such as nycodenz and metrizamide for their isolation.     

Characterization of neurite outgrowth and ectopic synaptogenesis in response to photoreceptor dysfunction

In the mammalian retina, light signals generated in photoreceptors are passed to bipolar and horizontal cells via synaptic contacts. In various pathological conditions, these second-order neurons extend neurites into the outer nuclear layer (ONL). However, the molecular events associated with this neurite outgrowth are not known. Here, we characterized the morphological synaptic changes in the CNGA3/CNGB1 double-knockout (A3B1) mouse, a model of retinitis pigmentosa. In these mice, horizontal cells looked normal until postnatal day (p) 11, but started growing neurites into the ONL 1 day later. At p28, the number of sprouting processes decreased, but the remaining sprouts developed synapse-like contacts at rod cell bodies, with an ultrastructural appearance reminiscent of ribbon synapses. Hence, neurite outgrowth and ectopic synaptogenesis in the A3B1 retina were precisely timed events starting at p12 and p28, respectively. We therefore performed microarray analysis of retinal gene expression in A3B1 and wild-type mice at those ages to evaluate the genomic response underlying these two events. This analysis identified 163 differentially regulated genes in the A3B1 retina related to neurite outgrowth or plasticity of synapses. The global changes in gene expression in the A3B1 retina were consistent with activation of signaling pathways related to Tp53, Smad, and Stat3. Moreover, key molecules of these signaling pathways could be localized at or in close proximity to outgrowing neurites. We therefore propose that Tp53, Smad, and Stat3 signaling pathways contribute to the synaptic plasticity in the A3B1 retina.     

Fine structure of modiffied phtoreceptor cells in the pineal of the gobyClevelandia ios (Pisces: Gobiidae)

Summary Photoreceptor cells with a system of long, thi cytoplasmic processes (50–80 nm thick), which amy represent a modification of the outer segment saccules, are described in the pineal of the goby,Clevelandia ios.The author wishes to thank Mr Larry Allen, who helped collect the specmens, and Mr Kyle Kikuchi, who was instrucmental in the preliminary studies.Partial support by Biomedical Research Support Grant (NIH) No. 447-12-000 from the Stritch School of Medicine is gratefully acknowledged.     

Morphological duality of the retinal pattern in flies.

In compound eyes of flies, a morphological duality of the retinal structure has been found. The retinal patterns based on this property and their possible evolutionary trends are described.     

Polo-box domain: a versatile mediator of polo-like kinase function

Members of the polo subfamily of protein kinases have emerged as important regulators in diverse aspects of the cell cycle and cell proliferation. A large body of evidence suggests that a highly conserved polo-box domain (PBD) present in the C-terminal non-catalytic region of polo kinases plays a pivotal role in the function of these enzymes. Recent advances in our comprehension of the mechanisms underlying mammalian polo-like kinase 1 (Plk1)-dependent protein–protein interactions revealed that the PBD serves as an essential molecular mediator that brings the kinase domain of Plk1 into proximity with its substrates, mainly through phospho-dependent interactions with its target proteins. In this review, current understanding of the structure and functions of PBD, mode of PBD-dependent interactions and substrate phosphorylation, and other phospho-independent functions of PBD are discussed.     

A note on the evolution of the transducer mechanism of the vertebrate retinal rod

Summary The Ca²⁺-gating mechanism that is the key component of membrane excitability in Paramecium has recently been localized on the cilia of this animal. Such a finding shows how the change in Ca²⁺ permeability (which is the probable consequence of the photoisomerization of rhodopsin) of the discs of the outer segments of the retinal rods could have evolved.