Defective Pigment Granule Biogenesis and Aberrant Behavior Caused by Mutations in the Drosophila AP-3β Adaptin Gene ruby

Abstract Lysosomal protein trafficking is a fundamental process conserved from yeast to humans. This conservation extends to lysosome-like organelles such as mammalian melanosomes and insect eye pigment granules. Recently, eye and coat color mutations in mouse (mocha and pearl) and Drosophila (garnet and carmine) were shown to affect subunits of the heterotetrameric adaptor protein complex AP-3 involved in vesicle trafficking. Here we demonstrate that the Drosophila eye color mutant ruby is defective in the AP-3β subunit gene. ruby expression was found in retinal pigment and photoreceptor cells and in the developing central nervous system. ruby mutations lead to a decreased number and altered size of pigment granules in various cell types in and adjacent to the retina. Humans with lesions in the related AP-3βA gene suffer from Hermansky-Pudlak syndrome, which is caused by defects in a number of lysosome-related organelles. Hermansky-Pudlak patients have a reduced skin pigmentation and suffer from internal bleeding, pulmonary fibrosis, and visual system malfunction. The Drosophila AP-3β adaptin also appears to be involved in processes other than eye pigment granule biogenesis because all ruby allele combinations tested exhibited defective behavior in a visual fixation paradigm.

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Vertebrate melanophores as potential model for drug discovery and development: A review

Cellular & Molecular Biology Letters ◽

10.2478/s11658-010-0044-y ◽

2011 ◽

Vol 16 (1) ◽

Cited By ~ 19

Author(s):

Saima Salim ◽

Sharique Ali

Keyword(s):

Drug Discovery ◽

High Throughput Screening ◽

Skin Pigmentation ◽

Physiological Mechanism ◽

Pigment Granule ◽

Pharmaceutical Products ◽

Surface Receptors ◽

Pigment Granules ◽

Intracellular Signals ◽

Specific Receptors

AbstractDrug discovery in skin pharmacotherapy is an enormous, continually expanding field. Researchers are developing novel and sensitive pharmaceutical products and drugs that target specific receptors to elicit concerted and appropriate responses. The pigment-bearing cells called melanophores have a significant contribution to make in this field. Melanophores, which contain the dark brown or black pigment melanin, constitute an important class of chromatophores. They are highly specialized in the bidirectional and coordinated translocation of pigment granules when given an appropriate stimulus. The pigment granules can be stimulated to undergo rapid dispersion throughout the melanophores, making the cell appear dark, or to aggregate at the center, making the cell appear light. The major signals involved in pigment transport within the melanophores are dependent on a special class of cell surface receptors called G-protein-coupled receptors (GPCRs). Many of these receptors of adrenaline, acetylcholine, histamine, serotonin, endothelin and melatonin have been found on melanophores. They are believed to have clinical relevance to skin-related ailments and therefore have become targets for high throughput screening projects. The selective screening of these receptors requires the recognition of particular ligands, agonists and antagonists and the characterization of their effects on pigment motility within the cells. The mechanism of skin pigmentation is incredibly intricate, but it would be a considerable step forward to unravel its underlying physiological mechanism. This would provide an experimental basis for new pharmacotherapies for dermatological anomalies. The discernible stimuli that can trigger a variety of intracellular signals affecting pigment granule movement primarily include neurotransmitters and hormones. This review focuses on the role of the hormone and neurotransmitter signals involved in pigment movement in terms of the pharmacology of the specific receptors.

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The occurrence of actinlike filaments in association with migrating pigment granules in frog retinal pigment epithelium

The Journal of Cell Biology ◽

10.1083/jcb.64.3.705 ◽

1975 ◽

Vol 64 (3) ◽

pp. 705-710 ◽

Cited By ~ 34

Author(s):

RL Murray ◽

MW Dubin

Keyword(s):

Pigment Epithelium ◽

Retinal Pigment ◽

Action Spectrum ◽

Pigment Granule ◽

Melanin Pigment ◽

Photoreceptor Outer Segments ◽

Outer Segments ◽

Pigment Granules ◽

Continuous Sheet ◽

Granule Motion

In the retina of the frog and certain other animals, melanin pigment granules move in response to light so as to shield photoreceptor outer segments. The granules are contained within the cells of the pigment epithelium (PE) which lie as a continuous sheet between the neural retina and the choroid. Moderate illumination of the eye causes the melanin granules to move from a region within a PE cell body into numerous fingerlike extensions of the cell which interdigitate with the receptor outer segments. This migration takes many minutes and is reversed when the light falling on the eye increases in intensity. Several reviews are concerned with the early descriptions of this phenomenon (6,30) and with more recent experiments (1,5,19). The mechanism of the pigment granule motion is undetermined although there are studies concerning PE ultrastructure (8, 23, 31), scanning electron microscopy of the fingerlike extensions of the PE cells (27), the role of the PE in photoreceptor phagocytosis (32), the nature of the pigment granules (19), and the action spectrum of the light which induces the migration (16). This study reports the presence of a system of microfilaments associated with the pigment granules in the fingerlike extensions processes of the PE cells. We demonstrate by heavy meromyosin (HMM) labeling that the filaments are actinlike in character and suggest that these filaments could be responsible for the migration of the melanin pigment granules.

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Interallelic Complementation at the Mouse Mitf Locus

Genetics ◽

10.1093/genetics/163.1.267 ◽

2003 ◽

Vol 163 (1) ◽

pp. 267-276

Author(s):

Eiríkur Steingrímsson ◽

Heinz Arnheiter ◽

Jón Hallsteinn Hallsson ◽

M Lynn Lamoreux ◽

Neal G Copeland ◽

...

Keyword(s):

Dna Binding ◽

Target Genes ◽

Coat Color ◽

Retinal Pigment ◽

Cell Types ◽

Binding Domain ◽

Retinal Pigment Epithelial Cells ◽

Dna Binding Domain ◽

Allelic Series ◽

Interallelic Complementation

Abstract Mutations at the mouse microphthalmia locus (Mitf) affect the development of different cell types, including melanocytes, retinal pigment epithelial cells of the eye, and osteoclasts. The MITF protein is a member of the MYC supergene family of basic-helix-loop-helix-leucine-zipper (bHLHZip) transcription factors and is known to regulate the expression of cell-specific target genes by binding DNA as homodimer or as heterodimer with related proteins. The many mutations isolated at the locus have different effects on the phenotype and can be arranged in an allelic series in which the phenotypes range from near normal to white microphthalmic animals with osteopetrosis. Previous investigations have shown that certain combinations of Mitf alleles complement each other, resulting in a phenotype more normal than that of each homozygote alone. Here we analyze this interallelic complementation in detail and show that it is limited to one particular allele, MitfMi-white (MitfMi-wh), a mutation affecting the DNA-binding domain. Both loss- and gain-of-function mutations are complemented, as are other Mitf mutations affecting the DNA-binding domain. Furthermore, this behavior is not restricted to particular cell types: Both eye development and coat color phenotypes are complemented. Our analysis suggests that MitfMi-wh-associated interallelic complementation is due to the unique biochemical nature of this mutation.

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Calcium-independent regulation of pigment granule aggregation and dispersion in teleost retinal pigment epithelial cells

Journal of Cell Science ◽

10.1242/jcs.109.1.33 ◽

1996 ◽

Vol 109 (1) ◽

pp. 33-43

Author(s):

C. King-Smith ◽

P. Chen ◽

D. Garcia ◽

H. Rey ◽

B. Burnside

Keyword(s):

Intracellular Calcium ◽

Pigment Epithelium ◽

Retinal Pigment ◽

Pigment Granule ◽

Retinal Pigment Epithelial Cells ◽

Rod Photoreceptor ◽

Rpe Cells ◽

Pigment Granules ◽

Pigment Granule Migration

In the eyes of teleosts and amphibians, melanin pigment granules of the retinal pigment epithelium (RPE) migrate in response to changes in light conditions. In the light, pigment granules disperse into the cells' long apical projections, thereby shielding the rod photoreceptor outer segments and reducing their extent of bleach. In darkness, pigment granules aggregate towards the base of the RPE cells. In vitro, RPE pigment granule aggregation can be induced by application of nonderivatized cAMP, and pigment granule dispersion can be induced by cAMP washout. In previous studies based on RPE-retina co-cultures, extracellular calcium was found to influence pigment granule migration. To examine the role of calcium in regulation of RPE pigment granule migration in the absence of retinal influences, we have used isolated RPE sheets and dissociated, cultured RPE cells. Under these conditions depletion of extracellular or intracellular calcium ([Ca2+]o, [Ca2+]i) had no effect on RPE pigment granule aggregation or dispersion. Using the intracellular calcium dye fura-2 and a new dye, fura-pe3, to monitor calcium dynamics in isolated RPE cells, we found that [Ca2+]i did not change from basal levels when pigment granule aggregation was triggered by cAMP, or dispersion was triggered by cAMP washout. Also, no change in [Ca2+]i was detected when dispersion was triggered by cAMP washout in the presence of 10 microM dopamine, a treatment previously shown to enhance dispersion. In addition, elevation of [Ca2+]i by addition of ionomycin neither triggered pigment movements, nor interfered with pigment granule motility elicited by cAMP addition or washout. Since other studies have indicated that actin plays a role in both pigment granule dispersion and aggregation in RPE, our findings suggest that RPE pigment granule migration depends on an actin-based motility system that is not directly regulated by calcium.

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The unusual microtubule polarity in teleost retinal pigment epithelial cells.

The Journal of Cell Biology ◽

10.1083/jcb.107.4.1461 ◽

1988 ◽

Vol 107 (4) ◽

pp. 1461-1464 ◽

Cited By ~ 29

Author(s):

L L Troutt ◽

B Burnside

Keyword(s):

Retinal Pigment Epithelial ◽

Pigment Epithelium ◽

Retinal Pigment ◽

Pigment Granule ◽

Cell Types ◽

Retinal Pigment Epithelial Cells ◽

Light Onset ◽

Rpe Cells ◽

Microtubule Polarity ◽

Pigment Epithelial Cells

In cells of the teleost retinal pigment epithelium (RPE), melanin granules disperse into the RPE cell's long apical projections in response to light onset, and aggregate toward the base of the RPE cell in response to dark onset. The RPE cells possess numerous microtubules, which in the apical projections are aligned longitudinally. Nocodazole studies have shown that pigment granule aggregation is microtubule-dependent (Troutt, L. L., and B. Burnside, 1988b Exp. Eye Res. In press.). To investigate further the mechanism of microtubule participation in RPE pigment granule aggregation, we have used the tubulin hook method to assess the polarity of microtubules in the apical projections of teleost RPE cells. We report here that virtually all microtubules in the RPE apical projections are uniformly oriented with plus ends toward the cell body and minus ends toward the projection tips. This orientation is opposite that found for microtubules of dermal melanophores, neurons, and most other cell types.

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Body Pigmentation as a Risk Factor for the Formation of Intracranial Aneurysms

BioMed Research International ◽

10.1155/2014/301631 ◽

2014 ◽

Vol 2014 ◽

pp. 1-9 ◽

Cited By ~ 1

Author(s):

Günter Schulter ◽

Klaus Leber ◽

Elke Kronawetter ◽

Viktoria R. Rübenbauer ◽

Peter Konstantiniuk ◽

...

Keyword(s):

Intracranial Aneurysms ◽

Coat Color ◽

Skin Pigmentation ◽

Pulmonary Veins ◽

Eye Color ◽

Age Related ◽

Brain Arteries ◽

Inflammatory Processes ◽

Eye Pigmentation ◽

Body Pigmentation

Recent studies demonstrated pigmented cells both in the murine heart, in pulmonary veins, and in brain arteries. Moreover, a role for melanocytes in the downregulation of inflammatory processes was suggested. As there is increasing evidence that inflammation is contributing significantly to the pathogenesis of intracranial aneurysms, melanocyte-like cells may be relevant in preventing age-related impairment of vessels. As pigmentation of the heart reflects that of coat color, aspects of body pigmentation might be associated with the incidence of intracranial aneurysms. We performed a case-control study to evaluate associations between the pigmentation of hair and eyes and the formation of aneurysms. In addition to hair and eye color, constitutive and facultative skin pigmentation were assessed in a replication study as well as individual handedness which can be seen as a neurophysiological correlate of developmental pigmentation processes. Hair pigmentation was highly associated with intracranial aneurysms in both samples, whereas eye pigmentation was not. In the replication cohort, facultative but not constitutive skin pigmentation proved significant. The strongest association was observed for individual handedness. Results indicate a significant association of intracranial aneurysms with particular aspects of body pigmentation as well as handedness, and imply clinical usefulness for screening of aneurysms and possible interventions.

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Protein kinase A regulation of pigment granule motility in retinal pigment epithelial cells from fish, Lepomis spp.

Visual Neuroscience ◽

10.1017/s0952523821000122 ◽

2021 ◽

Vol 38 ◽

Author(s):

Nicole E. Leitner ◽

Christina King-Smith

Keyword(s):

Protein Kinase ◽

Protein Kinase A ◽

Cell Body ◽

Single Cells ◽

Retinal Pigment ◽

Pigment Granule ◽

Mass Migration ◽

Pigment Granules ◽

Pka Pathway ◽

Kinase A

Abstract Retinomotor movements include elongation and contraction of rod and cone photoreceptors, and mass migration of melanin-containing pigment granules (melanosomes) of the retinal pigment epithelium (RPE) within the eyes of fish, frogs, and other lower vertebrates. Eyes of these animals do not contain dilatable pupils; therefore the repositioning of the rods and cones and a moveable curtain of pigment granules serve to modulate light intensity within the eye. RPE from sunfish (Lepomis spp.) can be isolated from the eye and dissociated into single cells, allowing in vitro studies of the cytoskeletal and regulatory mechanisms of organelle movement. Pigment granule aggregation from distal tips of apical projections into the cell body can be triggered by the application of underivatized cAMP, and dispersion is effected by cAMP washout in the presence of dopamine. While the phenomenon of cAMP-dependent pigment granule aggregation in isolated RPE was described many years ago, whether cAMP acts through the canonical cAMP-PKA pathway to stimulate motility has never been demonstrated. Here, we show that pharmacological inhibition of PKA blocks pigment granule aggregation, and microinjection of protein kinase A catalytic subunit triggers pigment granule aggregation. Treatment with a cAMP agonist that activates the Rap GEF, Epac (Effector protein activated by cAMP), had no effect on pigment granule position. Taken together, these results confirm that cAMP activates RPE pigment granule motility by the canonical cAMP-PKA pathway. Isolated RPE cells labeled with antibodies against PKA RIIα and against PKA-phosphorylated serine/threonine amino acids show diffuse, punctate labeling throughout the RPE cell body and apical projections. Immunoblotting of RPE lysates using the anti-PKA substrate antibody demonstrated seven prominent bands; two bands in particular at 27 and 64 kD showed increased levels of phosphorylation in the presence of cAMP, indicating their phosphorylation could contribute to the pigment granule aggregation mechanism.

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Ultrastructure of melanocyte-keratinocyte interactions and pigment transfer in vitro

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100084120 ◽

1978 ◽

Vol 36 (2) ◽

pp. 650-651

Author(s):

Raul I. Garcia ◽

Evelyn A. Flynn ◽

George Szabo

Keyword(s):

Direct Injection ◽

Skin Pigmentation ◽

Freeze Fracture ◽

Pigment Granule ◽

Time Lapse ◽

Ultrastructural Level ◽

Lanthanum Tracer ◽

A Cell

Skin pigmentation in mammals involves the interaction of epidermal melanocytes and keratinocytes in the structural and functional unit known as the Epidermal Melanin Unit. Melanocytes(M) synthesize melanin within specialized membrane-bound organelles, the melanosome or pigment granule. These are subsequently transferred by way of M dendrites to keratinocytes(K) by a mechanism still to be clearly defined. Three different, though not necessarily mutually exclusive, mechanisms of melanosome transfer have been proposed: cytophagocytosis by K of M dendrite tips containing melanosomes, direct injection of melanosomes into the K cytoplasm through a cell-to-cell pore or communicating channel formed by localized fusion of M and K cell membranes, release of melanosomes into the extracellular space(ECS) by exocytosis followed by K uptake using conventional phagocytosis. Variability in methods of transfer has been noted both in vivo and in vitro and there is evidence in support of each transfer mechanism. We Have previously studied M-K interactions in vitro using time-lapse cinemicrography and in vivo at the ultrastructural level using lanthanum tracer and freeze-fracture.

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The rosy locus in Drosophila melanogaster: xanthine dehydrogenase and eye pigments.

Genetics ◽

10.1093/genetics/129.4.1099 ◽

1991 ◽

Vol 129 (4) ◽

pp. 1099-1109 ◽

Cited By ~ 4

Author(s):

A G Reaume ◽

D A Knecht ◽

A Chovnick

Keyword(s):

Drosophila Melanogaster ◽

Structural Integrity ◽

Present Report ◽

Specific Interaction ◽

Pigment Granule ◽

Ultrastructural Localization ◽

Xanthine Dehydrogenase ◽

Pigment Formation ◽

Antibody Staining ◽

Pigment Granules

Abstract The rosy gene in Drosophila melanogaster codes for the enzyme xanthine dehydrogenase (XDH). Mutants that have no enzyme activity are characterized by a brownish eye color phenotype reflecting a deficiency in the red eye pigment. Xanthine dehydrogenase is not synthesized in the eye, but rather is transported there. The present report describes the ultrastructural localization of XDH in the Drosophila eye. Three lines of evidence are presented demonstrating that XDH is sequestered within specific vacuoles, the type II pigment granules. Histochemical and antibody staining of frozen sections, as well as thin layer chromatography studies of several adult genotypes serve to examine some of the factors and genic interactions that may be involved in transport of XDH, and in eye pigment formation. While a specific function for XDH in the synthesis of the red, pteridine eye pigments remains unknown, these studies present evidence that: (1) the incorporation of XDH into the pigment granules requires specific interaction between a normal XDH molecule and one or more transport proteins; (2) the structural integrity of the pigment granule itself is dependent upon the presence of a normal balance of eye pigments, a notion advanced earlier.

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