Publications by authors named "Steven G Britt"

15 Publications

  • Page 1 of 1

Drosophila R8 photoreceptor cell subtype specification requires hibris.

PLoS One 2020 14;15(10):e0240451. Epub 2020 Oct 14.

Department of Neurology, Department of Ophthalmology, Dell Medical School, University of Texas at Austin, Austin, Texas, United States of America.

Cell differentiation and cell fate determination in sensory systems are essential for stimulus discrimination and coding of environmental stimuli. Color vision is based on the differential color sensitivity of retinal photoreceptors, however the developmental programs that control photoreceptor cell differentiation and specify color sensitivity are poorly understood. In Drosophila melanogaster, there is evidence that the color sensitivity of different photoreceptors in the compound eye is regulated by inductive signals between cells, but the exact nature of these signals and how they are propagated remains unknown. We conducted a genetic screen to identify additional regulators of this process and identified a novel mutation in the hibris gene, which encodes an irre cell recognition module protein (IRM). These immunoglobulin super family cell adhesion molecules include human KIRREL and nephrin (NPHS1). hibris is expressed dynamically in the developing Drosophila melanogaster eye and loss-of-function mutations give rise to a diverse range of mutant phenotypes including disruption of the specification of R8 photoreceptor cell diversity. We demonstrate that hibris is required within the retina, and that hibris over-expression is sufficient to disrupt normal photoreceptor cell patterning. These findings suggest an additional layer of complexity in the signaling process that produces paired expression of opsin genes in adjacent R7 and R8 photoreceptor cells.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0240451PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7556441PMC
December 2020

Identification of Genes Involved in the Differentiation of R7y and R7p Photoreceptor Cells in .

G3 (Bethesda) 2020 11 5;10(11):3949-3958. Epub 2020 Nov 5.

Department of Neurology, Department of Ophthalmology, Dell Medical School, University of Texas at Austin, Austin, TX 78712

The R7 and R8 photoreceptor cells of the compound eye mediate color vision. Throughout the majority of the eye, these cells occur in two principal types of ommatidia. Approximately 35% of ommatidia are of the pale type and express in R7 cells and in R8 cells. The remaining 65% are of the yellow type and express in R7 cells and in R8 cells. The specification of an R8 cell in a pale or yellow ommatidium depends on the fate of the adjacent R7 cell. However, pale and yellow R7 cells are specified by a stochastic process that requires the genes , and To identify additional genes involved in this process we performed genetic screens using a collection of 480 transposon insertion strains. We identified genes in gain of function and loss of function screens that significantly altered the percentage of expressing R7 cells (%) from wild-type. 36 strains resulted in altered % in the gain of function screen where the insertion strains were crossed to a driver line. 53 strains resulted in altered % in the heterozygous loss of function screen. 4 strains showed effects that differed between the two screens, suggesting that the effect found in the gain of function screen was either larger than, or potentially masked by, the insertion alone. Analyses of homozygotes validated many of the candidates identified. These results suggest that R7 cell fate specification is sensitive to perturbations in mRNA transcription, splicing and localization, growth inhibition, post-translational protein modification, cleavage and secretion, signaling, ubiquitin protease activity, GTPase activation, actin and cytoskeletal regulation, and Ser/Thr kinase activity, among other diverse signaling and cell biological processes.
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http://dx.doi.org/10.1534/g3.120.401370DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7642934PMC
November 2020

The brachyceran de novo gene PIP82, a phosphorylation target of aPKC, is essential for proper formation and maintenance of the rhabdomeric photoreceptor apical domain in Drosophila.

PLoS Genet 2020 06 24;16(6):e1008890. Epub 2020 Jun 24.

Department of Biological Sciences, Wayne State University, Detroit, Michigan, United States of America.

The Drosophila apical photoreceptor membrane is defined by the presence of two distinct morphological regions, the microvilli-based rhabdomere and the stalk membrane. The subdivision of the apical membrane contributes to the geometrical positioning and the stereotypical morphology of the rhabdomeres in compound eyes with open rhabdoms and neural superposition. Here we describe the characterization of the photoreceptor specific protein PIP82. We found that PIP82's subcellular localization demarcates the rhabdomeric portion of the apical membrane. We further demonstrate that PIP82 is a phosphorylation target of aPKC. PIP82 localization is modulated by phosphorylation, and in vivo, the loss of the aPKC/Crumbs complex results in an expansion of the PIP82 localization domain. The absence of PIP82 in photoreceptors leads to misshapped rhabdomeres as a result of misdirected cellular trafficking of rhabdomere proteins. Comparative analyses reveal that PIP82 originated de novo in the lineage leading to brachyceran Diptera, which is also characterized by the transition from fused to open rhabdoms. Taken together, these findings define a novel factor that delineates and maintains a specific apical membrane domain, and offers new insights into the functional organization and evolutionary history of the Drosophila retina.
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http://dx.doi.org/10.1371/journal.pgen.1008890DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7340324PMC
June 2020

Chromophore-Independent Roles of Opsin Apoproteins in Drosophila Mechanoreceptors.

Curr Biol 2019 09 22;29(17):2961-2969.e4. Epub 2019 Aug 22.

Department of Cellular Neurobiology, University of Göttingen, 37077 Göttingen, Germany. Electronic address:

Rhodopsins, the major light-detecting molecules of animal visual systems [1], consist of opsin apoproteins that covalently bind a retinal chromophore with a conserved lysine residue [1, 2]. In addition to capturing photons, this chromophore contributes to rhodopsin maturation [3, 4], trafficking [3, 4], and stabilization [5], and defects in chromophore synthesis and recycling can cause dysfunction of the retina and dystrophy [6-9]. Indications that opsin apoproteins alone might have biological roles have come from archaebacteria and platyhelminths, which present opsin-like proteins that lack the chromophore binding site and are deemed to function independently of light [10, 11]. Light-independent sensory roles have been documented for Drosophila opsins [12-15], yet also these unconventional opsin functions are thought to require chromophore binding [12, 13, 15]. Unconjugated opsin apoproteins act as phospholipid scramblases in mammalian photoreceptor disks [16], yet chromophore-independent roles of opsin apoproteins outside of eyes have, to the best of our knowledge, hitherto not been described. Drosophila chordotonal mechanoreceptors require opsins [13, 15], and we find that their function remains uncompromised by nutrient carotenoid depletion. Disrupting carotenoid uptake and cleavage also left the mechanoreceptors unaffected, and manipulating the chromophore attachment site of the fly's major visual opsin Rh1 impaired photoreceptor, but not mechanoreceptor, function. Notwithstanding this chromophore independence, some proteins that process and recycle the chromophore in the retina are also required in mechanoreceptors, including visual cycle components that recycle the chromophore upon its photoisomerization. Our results thus establish biological function for unconjugated opsin apoproteins outside of eyes and, in addition, document chromophore-independent roles for chromophore pathway components.
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http://dx.doi.org/10.1016/j.cub.2019.07.036DOI Listing
September 2019

Analysis of Conserved Glutamate and Aspartate Residues in Drosophila Rhodopsin 1 and Their Influence on Spectral Tuning.

J Biol Chem 2015 Sep 20;290(36):21951-61. Epub 2015 Jul 20.

From the Departments of Cell and Developmental Biology, Ophthalmology and Rocky Mountain Lions Eye Institute, University of Colorado, Anschutz Medical Campus, School of Medicine, Aurora, Colorado 80045

The molecular mechanisms that regulate invertebrate visual pigment absorption are poorly understood. Studies of amphioxus Go-opsin have demonstrated that Glu-181 functions as the counterion in this pigment. This finding has led to the proposal that Glu-181 may function as the counterion in other invertebrate visual pigments as well. Here we describe a series of mutagenesis experiments to test this hypothesis and to also test whether other conserved acidic amino acids in Drosophila Rhodopsin 1 (Rh1) may serve as the counterion of this visual pigment. Of the 5 Glu and Asp residues replaced by Gln or Asn in our experiments, none of the mutant pigments shift the absorption of Rh1 by more than 6 nm. In combination with prior studies, these results suggest that the counterion in Drosophila Rh1 may not be located at Glu-181 as in amphioxus, or at Glu-113 as in bovine rhodopsin. Conversely, the extremely low steady state levels of the E194Q mutant pigment (bovine opsin site Glu-181), and the rhabdomere degeneration observed in flies expressing this mutant demonstrate that a negatively charged residue at this position is essential for normal rhodopsin function in vivo. This work also raises the possibility that another residue or physiologic anion may compensate for the missing counterion in the E194Q mutant.
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http://dx.doi.org/10.1074/jbc.M115.677765DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4571949PMC
September 2015

Phosphatidic acid phospholipase A1 mediates ER-Golgi transit of a family of G protein-coupled receptors.

J Cell Biol 2014 Jul;206(1):79-95

Laboratory of Cell and Developmental Signaling, National Cancer Institute, Frederick, MD 21702

The coat protein II (COPII)-coated vesicular system transports newly synthesized secretory and membrane proteins from the endoplasmic reticulum (ER) to the Golgi complex. Recruitment of cargo into COPII vesicles requires an interaction of COPII proteins either with the cargo molecules directly or with cargo receptors for anterograde trafficking. We show that cytosolic phosphatidic acid phospholipase A1 (PAPLA1) interacts with COPII protein family members and is required for the transport of Rh1 (rhodopsin 1), an N-glycosylated G protein-coupled receptor (GPCR), from the ER to the Golgi complex. In papla1 mutants, in the absence of transport to the Golgi, Rh1 is aberrantly glycosylated and is mislocalized. These defects lead to decreased levels of the protein and decreased sensitivity of the photoreceptors to light. Several GPCRs, including other rhodopsins and Bride of sevenless, are similarly affected. Our findings show that a cytosolic protein is necessary for transit of selective transmembrane receptor cargo by the COPII coat for anterograde trafficking.
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http://dx.doi.org/10.1083/jcb.201405020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4085702PMC
July 2014

Riding the crest of the wave: parallels between the neural crest and cancer in epithelial-to-mesenchymal transition and migration.

Wiley Interdiscip Rev Syst Biol Med 2013 Jul-Aug;5(4):511-22. Epub 2013 Apr 10.

Graduate Program in Cell Biology, Stem Cells and Development, School of Medicine, University of Colorado Anschutz Medical Campus, Aurora, CO, USA.

The neural crest (NC) is first induced as an epithelial population of cells at the neural plate border requiring complex signaling between bone morphogenetic protein, Wnt, and fibroblast growth factors to differentiate the neural and NC fate from the epidermis. Remarkably, following induction, these cells undergo an epithelial-to-mesenchymal transition (EMT), delaminate from the neural tube, and migrate through various tissue types and microenvironments before reaching their final destination where they undergo terminal differentiation. This process is mirrored in cancer metastasis, where a primary tumor will undergo an EMT before migrating and invading other cell populations to create a secondary tumor site. In recent years, as our understanding of NC EMT and migration has deepened, important new insights into tumorigenesis and metastasis have also been achieved. These discoveries have been driven by the observation that many cancers misregulate developmental genes to reacquire proliferative and migratory states. In this review, we examine how the NC provides an excellent model for studying EMT and migration. These data are discussed from the perspective of the gene regulatory networks that control both NC and cancer cell EMT and migration. Deciphering these processes in a comparative manner will expand our knowledge of the underlying etiology and pathogenesis of cancer and promote the development of novel targeted therapeutic strategies for cancer patients.
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http://dx.doi.org/10.1002/wsbm.1224DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3739939PMC
January 2014

Rhodopsin 5- and Rhodopsin 6-mediated clock synchronization in Drosophila melanogaster is independent of retinal phospholipase C-β signaling.

J Biol Rhythms 2012 Feb;27(1):25-36

School of Biological and Chemical Sciences, Queen Mary, University of London, London, United Kingdom.

Circadian clocks of most organisms are synchronized with the 24-hour solar day by the changes of light and dark. In Drosophila, both the visual photoreceptors in the compound eyes as well as the blue-light photoreceptor Cryptochrome expressed within the brain clock neurons contribute to this clock synchronization. A specialized photoreceptive structure located between the retina and the optic lobes, the Hofbauer-Buchner (H-B) eyelet, projects to the clock neurons in the brain and also participates in light synchronization. The compound eye photoreceptors and the H-B eyelet contain Rhodopsin photopigments, which activate the canonical invertebrate phototransduction cascade after being excited by light. We show here that 2 of the photopigments present in these photoreceptors, Rhodopsin 5 (Rh5) and Rhodopsin 6 (Rh6), contribute to light synchronization in a mutant (norpA(P41) ) that disrupts canonical phototransduction due to the absence of Phospholipase C-β (PLC-β). We reveal that norpA(P41) is a true loss-of-function allele, resulting in a truncated PLC-β protein that lacks the catalytic domain. Light reception mediated by Rh5 and Rh6 must therefore utilize either a different (nonretinal) PLC-β enzyme or alternative signaling mechanisms, at least in terms of clock-relevant photoreception. This novel signaling mode may distinguish Rhodopsin-mediated irradiance detection from image-forming vision in Drosophila.
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http://dx.doi.org/10.1177/0748730411431673DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4405110PMC
February 2012

Disruption of photoreceptor cell patterning in the Drosophila Scutoid mutant.

Fly (Austin) 2009 Oct-Dec;3(4):253-62. Epub 2009 Oct 7.

Department of Cell and Developmental Biology, University of Colorado Denver, School of Medicine, Aurora, CO, USA.

Cell fate determination in many systems is based upon inductive events driven by cell-cell interactions. Inductive signaling regulates many aspects of Drosophila compound eye development. Accumulating evidence suggests that the color sensitivity of the R8 photoreceptor cell within an individual ommatidium is regulated by an inductive signal from the adjacent R7 photoreceptor cell. This signal is thought to control an induced versus default cell-fate switch that coordinates the visual pigment expression and color sensitivities of adjacent R7 and R8 photoreceptor cells. Here we describe a disruption in R7 and R8 cell patterning in Scutoid mutants that is due to inappropriate signals from Rh4-expressing R7 cells inducing Rh5 expression in adjacent R8 cells. This dominant phenotype results from the misexpression of the transcriptional repressor snail, which with the co-repressor C-terminal-Binding-Protein represses rhomboid expression in the developing eye. We show that loss of rhomboid suppresses the Scutoid phenotype. However in contrast to the loss of rhomboid alone, which entirely blocks the normal inductive signal from the R7 to the R8 photoreceptor cell, Scutoid rhomboid double mutants display normal Rh5 and Rh6 expression. Our detailed analysis of this unusual dominant gain-of-function neomorphic phenotype suggests that the induction of Rh5 expression in Scutoid mutants is partially rhomboid independent.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2836898PMC
http://dx.doi.org/10.4161/fly.10546DOI Listing
March 2010

rhomboid mediates specification of blue- and green-sensitive R8 photoreceptor cells in Drosophila.

J Neurosci 2009 Mar;29(9):2666-75

Department of Cell and Developmental Biology, University of Colorado Denver, School of Medicine, Aurora, Colorado 80045, USA.

Color vision is based on the differential color sensitivity of retinal photoreceptors, however the developmental programs that control photoreceptor cell differentiation and specify color sensitivity are poorly understood. In Drosophila there is growing evidence that the color sensitivity of the R8 cell within an individual ommatidium is regulated by an inductive signal from the adjacent R7 cell. We previously examined the retinal patterning defect in Scutoid mutants, which results from a disruption of rhomboid expression. Here we show that loss of rhomboid blocks the induction of Rh5 expression and misexpression of rhomboid leads to the inappropriate induction of Rh5. These effects are specific to rhomboid, because its paralogue roughoid is neither required nor sufficient for the induction of Rh5 expression. We show that rhomboid is required cell-autonomously within the R8 photoreceptor cells and nonautonomously elsewhere in the eye for Rh5 induction. Interestingly, we found that the Epidermal growth factor receptor is also required for Rh5 induction, and its activation is sufficient to rescue the loss of Rh5 induction in a rhomboid mutant. This suggests that rhomboid may function in R8 cells to activate Epidermal growth factor receptor signaling in R7 cells and promote their differentiation to a signaling competent state.
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http://dx.doi.org/10.1523/JNEUROSCI.5988-08.2009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2679528PMC
March 2009

The green-absorbing Drosophila Rh6 visual pigment contains a blue-shifting amino acid substitution that is conserved in vertebrates.

J Biol Chem 2009 Feb 5;284(9):5717-22. Epub 2009 Jan 5.

Department of Cell and Developmental Biology, University of Colorado Denver, Aurora, Colorado 80045, USA.

The molecular mechanisms that regulate invertebrate visual pigment absorption are poorly understood. Through sequence analysis and functional investigation of vertebrate visual pigments, numerous amino acid substitutions important for this adaptive process have been identified. Here we describe a serine/alanine (S/A) substitution in long wavelength-absorbing Drosophila visual pigments that occurs at a site corresponding to Ala-292 in bovine rhodopsin. This S/A substitution accounts for a 10-17-nm absorption shift in visual pigments of this class. Additionally, we demonstrate that substitution of a cysteine at the same site, as occurs in the blue-absorbing Rh5 pigment, accounts for a 4-nm shift. Substitutions at this site are the first spectrally significant amino acid changes to be identified for invertebrate pigments sensitive to visible light and are the first evidence of a conserved tuning mechanism in vertebrate and invertebrate pigments of this class.
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http://dx.doi.org/10.1074/jbc.M807368200DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2645826PMC
February 2009

Two types of Drosophila R7 photoreceptor cells are arranged randomly: a model for stochastic cell-fate determination.

J Comp Neurol 2007 May;502(1):75-85

Department of Preventive and Social Medicine, University of Otago, Dunedin 9001, New Zealand.

The R7 photoreceptor cells of the Drosophila retina are ultraviolet sensitive and are thought to mediate color discrimination and polarized light detection. In addition, there is growing evidence that the color sensitivity of the R8 cell within an individual ommatidium is regulated by a genetic switch that depends on the type of R7 cell adjacent to it. Here we examine the organization of the two major types of R7 cells by three different rigorous statistical methods and present evidence that they are arranged randomly and independently. First, we performed L-function analyses to test whether the organization of R7 cells (and the relationship between them) is regular, clustered, or completely spatially random. Next, we used generalized linear mixed models to test whether the proportion of R7 cell neighbors differs from their prevalence within the eye as a whole. Finally, we conducted a series of simulations to test whether the proportion of R7 cell neighbors differs from that in a random simulation. In each case, we found evidence that the organization of the two types of R7 cells is random and independent, suggesting that R7 cells in neighboring ommatidia are unlikely to interact and influence each other's identity and may be determined stochastically in a cell-autonomous manner. Compared with traditional lineage or inductive mechanisms, this may represent a novel mechanism of cell fate determination based on noisy or stochastic gene expression in which the differentiation of an individual R7 cell is a random event but the proportions of R7 cell subtypes are regulated.
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http://dx.doi.org/10.1002/cne.21298DOI Listing
May 2007

Expression of Drosophila rhodopsins during photoreceptor cell differentiation: insights into R7 and R8 cell subtype commitment.

Gene Expr Patterns 2006 Oct 21;6(7):687-94. Epub 2006 Feb 21.

Department of Cell and Developmental Biology, University of Colorado at Denver and Health Sciences Center, School of Medicine, RC1 South, Aurora, 80045, USA.

The R7 and R8 photoreceptor cells of the Drosophila retina are thought to mediate color discrimination and polarized light detection. This is based on the patterned expression of different visual pigments, rhodopsins, in different photoreceptor cells. In this report, we examined the developmental timing of retinal patterning. There is genetic evidence that over the majority of the eye, patterned expression of opsin genes is regulated by a signal from one subtype of R7 cells to adjacent R8 cells. We examined the onset of expression of the rhodopsin genes to determine the latest time point by which photoreceptor subtype commitment must have occurred. We found that the onset of rhodopsin expression in all photoreceptors of the compound eye occurs during a narrow window from 79% to 84% of pupal development (approximately 8 h), pupal stages P12-P14. Rhodopsin 1 has the earliest onset, followed by Rhodopsins 3, 4, and 5 at approximately the same time, and finally Rhodopsin 6. This sequence mimics the model for how R7 and R8 photoreceptor cells are specified, and defines the timing of photoreceptor cell fate decisions with respect to other events in eye development.
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http://dx.doi.org/10.1016/j.modgep.2006.01.003DOI Listing
October 2006

Molecular basis for ultraviolet vision in invertebrates.

J Neurosci 2003 Nov;23(34):10873-8

Department of Cell and Developmental Biology, University of Colorado Health Sciences Center, Denver, Colorado 80262, USA.

Invertebrates are sensitive to a broad spectrum of light that ranges from UV to red. Color sensitivity in the UV plays an important role in foraging, navigation, and mate selection in both flying and terrestrial invertebrate animals. Here, we show that a single amino acid polymorphism is responsible for invertebrate UV vision. This residue (UV: lysine vs blue:asparagine or glutamate) corresponds to amino acid position glycine 90 (G90) in bovine rhodopsin, a site affected in autosomal dominant human congenital night blindness. Introduction of the positively charged lysine in invertebrates is likely to deprotonate the Schiff base chromophore and produce an UV visual pigment. This same position is responsible for regulating UV versus blue sensitivity in several bird species, suggesting that UV vision has arisen independently in invertebrate and vertebrate lineages by a similar molecular mechanism.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2819302PMC
November 2003

Heterologous expression of limulus rhodopsin.

J Biol Chem 2003 Oct 23;278(42):40493-502. Epub 2003 Jun 23.

Department of Ophthalmology, SUNY Upstate Medical University, Syracuse, New York 13210, USA.

Invertebrates such as Drosophila or Limulus assemble their visual pigment into the specialized rhabdomeric membranes of photoreceptors where phototransduction occurs. We have investigated the biosynthesis of rhodopsin from the Limulus lateral eye with three cell culture expression systems: mammalian COS1 cells, insect Sf9 cells, and amphibian Xenopus oocytes. We extracted and affinity-purified epitope-tagged Limulus rhodopsin expressed from a cDNA or cRNA from these systems. We found that all three culture systems could efficiently synthesize the opsin polypeptide in quantities comparable with that found for bovine opsin. However, none of the systems expressed a protein that stably bound 11-cis-retinal. The protein expressed in COS1 and Sf9 cells appeared to be misfolded, improperly localized, and proteolytically degraded. Similarly, Xenopus oocytes injected with Limulus opsin cRNA did not evoke light-sensitive currents after incubation with 11-cis-retinal. However, injecting Xenopus oocytes with mRNA from Limulus lateral eyes yielded light-dependent conductance changes after incubation with 11-cis-retinal. Also, expressing Limulus opsin cDNA in the R1-R6 photoreceptors of transgenic Drosophila yielded a visual pigment that bound retinal, had normal spectral properties, and coupled to the endogenous phototransduction cascade. These results indicate that Limulus opsin may require one or more photoreceptor-specific proteins for correct folding and/or chromophore binding. This may be a general property of invertebrate opsins and may underlie some of the functional differences between invertebrate and vertebrate visual pigments.
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http://dx.doi.org/10.1074/jbc.M304567200DOI Listing
October 2003