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Nobuhiro Nagai, Satoru Iwata, Hirokazu Kaji, Kaori Sampei, Yuki Katsukura, Hideyuki Onami, Matsuhiko Nishizawa, Toru Nakazawa, Yukihiko Mashima, Toshiaki Abe
It has been suggested that unoprostone isopropyl (UNO) has potent neuroprotective activity in the retina. The effect of sustained transscleral UNO delivery to the posterior segment of the eye on photoreceptor degeneration was evaluated. UNO was loaded into a device made of poly(ethyleneglycol) dimethacrylate by polydimethylsiloxane mold-based UV-curing. The amount of UNO diffusing from these devices was measured using high-performance liquid chromatography. The polymeric devices that released UNO at 1.8 μg/day were implanted on the sclerae of S334ter rats at postnatal 21 days, and electroretinograms (ERGs) were compared with those of topical application and placebo devices...
November 2016: Journal of Biomedical Materials Research. Part B, Applied Biomaterials
Tapasi Rana, Pravallika Kotla, Roderick Fullard, Marina Gorbatyuk
Expression of T17M rhodopsin (T17M) in rods activates the Unfolded Protein Response (UPR) and leads to the development of autosomal dominant retinitis pigmentosa (adRP). The rod death occurs in adRP retinas prior to cone photoreceptor death, so the mechanism by which cone photoreceptors die remains unclear. Therefore, the goal of the study was to verify whether UPR in rods induces TNFa-mediated signaling to the cones and to determine whether the TNFa deficit could prevent adRP cone cell death. Primary rod photoreceptors and cone-derived 661W cells transfected with siRNA against TNFa were treated with tunicamycin to mimic activation of UPR in T17M retinas expressing normal and reduced TNFa levels...
October 14, 2016: Biochimica et Biophysica Acta
Anna Ngo, Ann Koay, Christian Pecquet, Carmen C Diaconu, Yasmine Ould-Amer, Qiwei Huang, Congbao Kang, Anders Poulsen, May Ann Lee, David Jenkins, Andrew Shiau, Stefan N Constantinescu, Meng Ling Choo
BACKGROUND: Rather than a Janus Kinase 2 inhibitor (ruxolitinib), a specific thrombopoietin receptor (TpoR) inhibitor would be more specific for the treatment of myeloproliferative neoplasms due to TpoR mutations. OBJECTIVE: A cell-based phenotypic approach to identify specific TpoR inhibitors was implemented and a library of 505,483 small molecules was screened for inhibitory effects on cells transformed by TpoR mutants. RESULTS: Among the identified hits are two analogs of 3-(4-piperidinyl) indole...
October 10, 2016: Combinatorial Chemistry & High Throughput Screening
Tatiana B Feldman, Olga A Smitienko, Ivan V Shelaev, Fedor E Gostev, Oksana V Nekrasova, Dmitriy A Dolgikh, Victor A Nadtochenko, Mikhail P Kirpichnikov, Mikhail A Ostrovsky
Photochromic ultrafast reactions of bacteriorhodopsin (H. salinarum) and bovine rhodopsin were conducted with a femtosecond two-pump probe pulse setup with the time resolution of 20-25fs. The dynamics of the forward and reverse photochemical reactions for both retinal-containing proteins was compared. It is demonstrated that when retinal-containing proteins are excited by femtosecond pulses, dynamics pattern of the vibrational coherent wave packets in the course of the reaction is different for bacteriorhodopsin and visual rhodopsin...
October 2, 2016: Journal of Photochemistry and Photobiology. B, Biology
Suchithranga M D C Perera, Udeep Chawla, Michael F Brown
Preparation and storage of functional membrane proteins such as G-protein-coupled receptors (GPCRs) are crucial to the processes of drug delivery and discovery. Here we describe a method of preparing powdered GPCRs using rhodopsin as the prototype. We purified rhodopsin in CHAPS detergent with low detergent to protein ratio so the bulk of the sample represented protein (ca. 72% w/w). Our new method for generating powders of membrane proteins followed by rehydration paves the way for conducting functional and biophysical experiments...
October 10, 2016: Journal of Physical Chemistry Letters
Takaaki Sokabe, Hsiang-Chin Chen, Junjie Luo, Craig Montell
Drosophila third-instar larvae exhibit changes in their behavioral responses to gravity and food as they transition from feeding to wandering stages. Using a thermal gradient encompassing the comfortable range (18°C to 28°C), we found that third-instar larvae exhibit a dramatic shift in thermal preference. Early third-instar larvae prefer 24°C, which switches to increasingly stronger biases for 18°C-19°C in mid- and late-third-instar larvae. Mutations eliminating either of two rhodopsins, Rh5 and Rh6, wiped out these age-dependent changes in thermal preference...
October 4, 2016: Cell Reports
Reeshan Ul Quraish, Norihiro Sudou, Kaori Nomura-Komoike, Fumi Sato, Hiroki Fujieda
PURPOSE: p27(KIP1) (p27), originally identified as a cell cycle inhibitor, is now known to have multifaceted roles beyond cell cycle regulation. p27 is required for the normal histogenesis of the RPE, but the role of p27 in the mature RPE remains elusive. To define the role of p27 in the maintenance and function of the RPE, we investigated the effects of p27 deletion on the responses of the RPE after photoreceptor damage. METHODS: Photoreceptor damage was induced in wild-type (WT) and p27 knockout (KO) mice with N-methyl-N-nitrosourea (MNU) treatment...
2016: Molecular Vision
Christopher T Schafer, Jonathan F Fay, Jay M Janz, David L Farrens
Here, we describe two insights into the role of receptor conformational dynamics during agonist release (all-trans retinal, ATR) from the visual G protein-coupled receptor (GPCR) rhodopsin. First, we show that, after light activation, ATR can continually release and rebind to any receptor remaining in an active-like conformation. As with other GPCRs, we observe that this equilibrium can be shifted by either promoting the active-like population or increasing the agonist concentration. Second, we find that during decay of the signaling state an active-like, yet empty, receptor conformation can transiently persist after retinal release, before the receptor ultimately collapses into an inactive conformation...
October 4, 2016: Proceedings of the National Academy of Sciences of the United States of America
Xiao Wang, Yawen Mu, Mengshi Sun, Junhai Han
Homeostatic regulation of the light sensor, rhodopsin, is critical for the maintenance of light sensitivity and survival of photoreceptors. The major fly rhodopsin, Rh1, undergoes light-induced endocytosis and degradation, but its protein and mRNA levels remain constant during light/dark cycles. It is not clear how translation of Rh1 is regulated. Here, we show that adult photoreceptors maintain a constant, abundant quantity of ninaE mRNA, which encodes Rh1. We demonstrate that the Fmr1 protein associates with ninaE mRNA and represses its translation...
October 4, 2016: Journal of Molecular Cell Biology
Mayanka Awasthi, Peeyush Ranjan, Komal Sharma, Sindhu Kandoth Veetil, Suneel Kateriya
The bacterial type rhodopsins are present in all the three domains of life. In contrast to the animal type rhodopsin that performs mainly sensory functions in higher eukaryotes, the bacterial type rhodopsin could function as ion channel, pumps and as sensory proteins. The functioning of rhodopsin in higher eukaryotes requires the transport of rhodopsin from its site of synthesis to the ciliated outer segment of the photoreceptive cells. However, the trafficking of bacterial type rhodopsin from its site of synthesis to the position of action is not characterized...
October 3, 2016: Scientific Reports
Birgit Ploier, Lydia N Caro, Takefumi Morizumi, Kalpana Pandey, Jillian N Pearring, Michael A Goren, Silvia C Finnemann, Johannes Graumann, Vadim Y Arshavsky, Jeremy S Dittman, Oliver P Ernst, Anant K Menon
Retinitis pigmentosa (RP) is a blinding disease often associated with mutations in rhodopsin, a light-sensing G protein-coupled receptor and phospholipid scramblase. Most RP-associated mutations affect rhodopsin's activity or transport to disc membranes. Intriguingly, some mutations produce apparently normal rhodopsins that nevertheless cause disease. Here we show that three such enigmatic mutations-F45L, V209M and F220C-yield fully functional visual pigments that bind the 11-cis retinal chromophore, activate the G protein transducin, traffic to the light-sensitive photoreceptor compartment and scramble phospholipids...
October 3, 2016: Nature Communications
Birgit Ploier, Anant K Menon
Scramblases translocate phospholipids across the membrane bilayer bidirectionally in an ATP-independent manner. The first scramblase to be identified and biochemically verified was opsin, the apoprotein of the photoreceptor rhodopsin. Rhodopsin is a G protein-coupled receptor localized in rod photoreceptor disc membranes of the retina where it is responsible for the perception of light. Rhodopsin's scramblase activity does not depend on its ligand 11-cis-retinal, i.e., the apoprotein opsin is also active as a scramblase...
2016: Journal of Visualized Experiments: JoVE
E G Govorunova, L A Koppel
Optogenetics technology (using light-sensitive microbial proteins to control animal cell physiology) is becoming increasingly popular in laboratories around the world. Among these proteins, particularly important are rhodopsins that transport ions across the membrane and are used in optogenetics to regulate membrane potential by light, mostly in neurons. Although rhodopsin ion pumps transport only one charge per captured photon, channelrhodopsins are capable of more efficient passive transport. In this review, we follow the history of channelrhodopsin discovery in flagellate algae and discuss the latest addition to the channelrhodopsin family, channels with anion, rather than cation, selectivity...
September 2016: Biochemistry. Biokhimii︠a︡
Rei Abe-Yoshizumi, Keiichi Inoue, Hideaki E Kato, Osamu Nureki, Hideki Kandori
Light-driven outward sodium-pumping rhodopsin (NaR) was recently found in marine bacteria. Krokinobacter eikastus rhodopsin 2 (KR2) actively transports sodium and lithium ions in NaCl and LiCl, respectively, while it pumps protons in KCl. NaR has a conserved NDQ (N112, D116, and Q123 in KR2) motif, and previous studies suggested an important role for N112 in the function of KR2. Here we replaced N112 with 19 different amino acids and studied the molecular properties of the mutants. All mutants exhibited absorption bands from a protonated Schiff base in the λmax range from 508 to 531 nm upon heterologous expression in Escherichia coli, whose ion-pumping activity was measured using pH electrodes...
October 6, 2016: Biochemistry
Prashant Kumar, Anisha Ashokan, Gopala Krishna Aradhyam
BACKGROUND: Human APJ receptor (APJR), a rhodopsin family G-Protein Coupled Receptor (GPCR), activated by isoforms of peptide ligand apelin causing potent inotropic effect, is involved in cardiac function, angiogenesis and maintenance of fluid homeostasis. APJR is expressed in various organs e.g., heart, brain, kidney, muscles, etc. Hence, problems in APJR signaling lead to severe dysregulation in the pathophysiology of an organism. METHODS: Based on multiple sequence alignment of receptors from various organisms, we observe a large number of conserved residues in the extracellular side...
September 23, 2016: Biochimica et Biophysica Acta
Ralf Thiel, Thomas Knebelsberger
Accurate stock assessments for each of the dominant species of sand lances in the northeast Atlantic Ocean and adjacent areas are not available due to the lack of a reliable identification procedure; therefore, appropriate measures of fisheries management or conservation of sand lances cannot be implemented. In this study, detailed morphological and molecular features are assessed to discriminate between four species of sand lances belonging to the genera Ammodytes and Hyperoplus. Morphological characters described by earlier authors as useful for identification of the genera are confirmed, and two additional distinguishing characters are added...
2016: ZooKeys
Lan-Ping Zheng, Xiao-Yong Chen, Jun-Xing Yang
The major phylogenetic pattern of the cyprinid tribe Labeonini has been revealed by previous molecular studies; however, the relationships within a clade that mainly inhabits the karst regions, which we refer to as the "karst group", in southwest China remain unresolved due to the low taxon sampling. This group includes more than 50% of the genera and species of Labeonini in China. Moreover, more than 90% of the genera of this group are endemic to China. In addition, some new genera and species of Labeonini have been discovered from these karst regions, but their taxonomic validity and phylogenetic position have not been examined...
2016: ZooKeys
Sara M Duncan, Gail M Seigel
Historically, standard enzyme immunohistochemistry has been accomplished with brown (DAB, diaminobenzidine) substrate. This can become problematic in pigmented tissues, such as the retina, where brown pigment of retinal pigmented epithelial (RPE) cells can be easily confounded with brown substrate. Although immunofluorescence detection methods can overcome this challenge, fluorescence may fade over a period of weeks, while enzyme substrates allow for more long-lasting, archival results. In this report, we will describe a high-contrast enzyme immunohistochemistry method ideal for pigmented tissues that utilizes purple (VIP) substrate...
2016: Journal of Biological Methods
Anne C Roulin, Yann Bourgeois, Urs Stiefel, Jean-Claude Walser, Dieter Ebert
Diapause is an adaptation that allows organisms to survive harsh environmental conditions. In species occurring over broad habitat ranges, both the timing and the intensity of diapause induction can vary across populations, revealing patterns of local adaptation. Understanding the genetic architecture of this fitness-related trait would help clarify how populations adapt to their local environments. In the cyclical parthenogenetic crustacean Daphnia magna, diapause induction is a phenotypic plastic life history trait linked to sexual reproduction, as asexual females have the ability to switch to sexual reproduction and produce resting stages, their sole strategy for surviving habitat deterioration...
September 22, 2016: Molecular Biology and Evolution
Shintaro Nakamura, Takashi Kikukawa, Jun Tamogami, Masakatsu Kamiya, Tomoyasu Aizawa, Martin W Hahn, Kunio Ihara, Naoki Kamo, Makoto Demura
Actinorhodopsin (ActR) is a light-driven outward H(+) pump. Although the genes of ActRs are widely spread among freshwater bacterioplankton, there are no prior data on their functional expression in native cell membranes. Here, we demonstrate ActR phototrophy in the native actinobacterium. Genome analysis showed that Candidatus Rhodoluna planktonica, a freshwater actinobacterium, encodes one microbial rhodopsin (RpActR) belonging to the ActR family. Reflecting the functional expression of RpActR, illumination induced the acidification of the actinobacterial cell suspension and then elevated the ATP content inside the cells...
September 20, 2016: Biochimica et Biophysica Acta
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