The development of melatonin-binding sites in the ovine fetus

1994 ◽  
Vol 142 (3) ◽  
pp. 475-484 ◽  
Author(s):  
R J A Helliwell ◽  
L M Williams
Keyword(s):  
G Protein ◽  
Binding Sites ◽  
Early Stage ◽  
Specific Binding ◽  
Melatonin Receptors ◽  
Pars Tuberalis ◽  
Dependent Manner ◽  
Pars Distalis ◽  
Specific Expression ◽  
Fetal Sheep

Abstract The pineal hormone, melatonin, is important in the timing of seasonal reproduction in the sheep. Melatonin of maternal origin readily crosses the placenta; its function in the fetal sheep is, however, unclear. To gain an insight into the role of melatonin in ovine development we have identified specific melatonin receptors throughout gestation using 2-[125I]iodomelatonin and quantitative in vitro autoradiography. Specific binding was found at the earliest time studied at 30 days of gestation, over the developing thyroid (term=145 days). At 31 days of gestation specific labelling was found over the thyroid and pituitary glands, the spinal nerves, nasal cavity and developing bronchi. This binding was diminished by over 50% in the presence of 10−4 m GTPγS (an analogue of guanosine triphosphate) indicating that the 2-[125I]iodomelatonin binding at this early stage of gestation represents a receptor coupled to a regulatory G-protein. By 40 days of gestation specific binding was found over the nasal epithelium, cochlear epithelium, regions of the brain, especially the hind brain and the vestibulocochlear and glossopharyngeal nerves, and both the pars distalis and pars tuberalis of the pituitary. As gestation proceeded, labelling over the pars distalis appeared to become more scattered in nature while that on the pars tuberalis remained consistent. Saturation studies of both the neuronal and pituitary binding sites at 121 days of gestation and in the newborn lamb revealed a single class of high-affinity binding sites with Kd values in the picomolar range. Also at 121 days of gestation, binding over the fetal pars tuberalis was diminished in a dose-dependent manner by GTPγS, again confirming that specific binding is indicative of a receptor coupled to a regulatory G-protein. These data demonstrate a potential for sensitivity to melatonin from early in gestation, as well as the developmentally specific expression of the melatonin receptor in certain tissues, and suggest a wider role for melatonin in ovine fetal development than previously considered. Journal of Endocrinology (1994) 142, 475–484

A carnivore species (Canis familiaris) expresses circadian melatonin rhythm in the peripheral blood and melatonin receptors in the brain

1994 ◽  
Vol 131 (2) ◽  
pp. 191-200 ◽  
Author(s):  
Bojidar Stankov ◽  
Morten Møller ◽  
Valeria Lucini ◽  
Simona Capsoni ◽  
Franco Fraschini
Keyword(s):  
Peripheral Blood ◽  
Binding Sites ◽  
Specific Binding ◽  
Canis Familiaris ◽  
Melatonin Receptors ◽  
Pars Tuberalis ◽  
Pars Distalis ◽  
Melatonin Rhythm ◽  
Carnivore Species ◽  
The Brain

Stankov B, Møller, M, Lucini V, Capsoni S, Fraschini F. A carnivore species (Canis familiaris) expresses circadian melatonin rhythm in the peripheral blood and melatonin receptors in the brain. Eur J Endocrinol 1994;131:191–200. ISSN 0804–4643 Dogs kept under controlled photoperiodic conditions of 12 h light and 12 h dark expressed a clear diurnal melatonin rhythm in the peripheral blood, with a swift peak restricted to the late part of the scotophase. The highest density of high-affinity, G-protein-linked 2-[125I]iodomelatonin binding sites was found in the pars tuberalis of the pituitary gland. Binding sites were found also in the pars distalis, and light microscopy/high-resolution autoradiography showed that binding was located exclusively over the chromophobe and basophilic cells forming the adenopituitary zona tuberalis, well developed in this species, and extending into the gland as a continuation of pars tuberalis. Cords of basophilic cells located in the pars distalis proper also expressed high receptor density. The eosinophils in the adenohypophysis and the neural lobe were devoid of binding. Heavily labeled were the external laminar and the mitral cell layers of the olfactory bulbs, but no binding was detected in the filae nervi olfactorii or tractus olfactorius. The hypothalamic suprachiasmatic nuclei were discernible clearly. Quantitative autoradiography inhibition experiments revealed that the apparent melatonin inhibitory constant (ic50) in all those areas was around 0.1 nmol/l, which is a physiologically appropriate value considering the peripheral blood melatonin levels. Co-incubation with guanosine 5′-O-(3-thiotriphosphate) (GTPΓS) led to a consequential decrease in the binding density. The specific binding observed in other areas (hippocampus, frontal, parietal, occipital cortex and cerebellum) was rather weak, diffuse and could not be attributed to a particular layer; the apparent ic50 for melatonin was about 1 μmol/l, and co-incubation with GTPΓS did not modify the binding density. Collectively, these data show that the dog posesses all the prerequisites for an efficient network adapted to photoperiodic time measurements. A circadian melatonin signal in the peripheral blood and an apparently functional readout receptor system located in key positions within the brain are both present in this species. Bojidar Stankov, Chair of Chemotherapy, Department of Pharmacology, University of Milan, 32 Via Vanvitelli, 20129 Milano, Italy

Melatonin-binding sites in the rat brain and pituitary mapped by in-vitro autoradiography

1989 ◽  
Vol 3 (1) ◽  
pp. 71-75 ◽  
Author(s):  
L. M. Williams
Keyword(s):  
Rat Brain ◽  
Binding Sites ◽  
Area Postrema ◽  
Specific Binding ◽  
Melatonin Receptors ◽  
Pars Tuberalis ◽  
Lower Affinity ◽  
High Affinity ◽  
In Vitro Autoradiography

ABSTRACT Using picomolar concentrations of [125I]iodomelatonin and in-vitro autoradiography, specific melatonin-binding sites have been mapped in the rat brain and pituitary. Using this same technique, high-affinity melatonin receptors had previously been identified in the suprachiasmatic nucleus (SCN) and median eminence regions of the rat hypothalamus. The presence of melatonin binding in the SCN has been confirmed, but the second area of binding has been identified as the pars tuberalis of the pituitary, and a completely novel area of binding is also reported in the area postrema. The existence of lower affinity melatonin receptors in the rat brain was also investigated using in-vitro autoradiography and higher concentrations of [125I]iodomelatonin. No further sites of specific binding were, however, disclosed.

scholarly journals Mot3, a Zn Finger Transcription Factor That Modulates Gene Expression and Attenuates Mating Pheromone Signaling in Saccharomyces cerevisiae

Genetics ◽  
1998 ◽  
Vol 149 (2) ◽  
pp. 879-892 ◽  
Author(s):  
Anatoly V Grishin ◽  
Michael Rothenberg ◽  
Maureen A Downs ◽  
Kendall J Blumer
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Signaling Pathway ◽  
G Protein ◽  
Binding Sites ◽  
Dependent Manner ◽  
Pheromone Response ◽  
Mating Pheromone ◽  
Pheromone Signaling ◽  
Yeast Genes

Abstract In the yeast Saccharomyces cerevisiae, mating pheromone response is initiated by activation of a G protein- and mitogen-activated protein (MAP) kinase-dependent signaling pathway and attenuated by several mechanisms that promote adaptation or desensitization. To identify genes whose products negatively regulate pheromone signaling, we screened for mutations that suppress the hyperadaptive phenotype of wild-type cells overexpressing signaling-defective G protein β subunits. This identified recessive mutations in MOT3, which encodes a nuclear protein with two Cys2-His2 Zn fingers. MOT3 was found to be a dosage-dependent inhibitor of pheromone response and pheromone-induced gene expression and to require an intact signaling pathway to exert its effects. Several results suggested that Mot3 attenuates expression of pheromone-responsive genes by mechanisms distinct from those used by the negative transcriptional regulators Cdc36, Cdc39, and Mot2. First, a Mot3-lexA fusion functions as a transcriptional activator. Second, Mot3 is a dose-dependent activator of several genes unrelated to pheromone response, including CYC1, SUC2, and LEU2. Third, insertion of consensus Mot3 binding sites (C/A/T)AGG(T/C)A activates a promoter in a MOT3-dependent manner. These findings, and the fact that consensus binding sites are found in the 5′ flanking regions of many yeast genes, suggest that Mot3 is a globally acting transcriptional regulator. We hypothesize that Mot3 regulates expression of factors that attenuate signaling by the pheromone response pathway.

Atrial natriuretic peptide in the eel, Anguilla anguilla L.: its cardiac distribution, receptors and actions on isolated branchial cells

1992 ◽  
Vol 9 (2) ◽  
pp. 103-114 ◽  
Author(s):  
C. L. Broadhead ◽  
U. T. O'Sullivan ◽  
C. F. Deacon ◽  
I. W. Henderson
Keyword(s):  
Sodium Chloride ◽  
Atrial Natriuretic Peptide ◽  
Natriuretic Peptide ◽  
Binding Sites ◽  
Specific Binding ◽  
Anguilla Anguilla ◽  
Chloride Cells ◽  
Dependent Manner ◽  
Single Class ◽  
Atrial Natriuretic

ABSTRACT The presence of atrial natriuretic peptide (ANP) and the nature of its binding sites were studied in freshwater (FW)- and seawater (SW)-adapted eels using a heterologous analogue, that of the rat (rANP). Rat ANP-like immunoreactivity was demonstrated in the cardiac atria and ventricles of both FW and SW eels, and electron-dense ANP-like granules were observed. The atria and ventricles of FW eels contained significantly more granules than those of SW animals and, in both types, the atria were more granular than the ventricles. Specific binding sites for rANP were demonstrated by displacement and uptake experiments using labelled rANP in dispersed eel branchial cell preparations, enriched in chloride cells. The concentration of rANP required to produce a 50% inhibition of binding in FW cells was significantly lower than that in SW cells. Scatchard analyses revealed the presence of two classes of binding site in SW eel branchial cells but only a single class of receptor in FW cells. The affinity of the FW receptor was not significantly different from that of the SW high affinity site. Rat ANP stimulated the production of cyclic GMP (cGMP) in a dose-dependent manner, and both basal and stimulated levels of cGMP were significantly greater in SW branchial cells. These studies suggest that ANP is involved in the adaptation of the euryhaline eel to differing environmental salinities; the levels of the peptide in the heart alter with changing salinity, and the nature of the receptors in the sodium chloride-transporting epithelium of the gill changes in response to the need either to eliminate or to absorb sodium chloride.

Characterization of melatonin receptors in the ram pars tuberalis: influence of light

1990 ◽  
Vol 123 (5) ◽  
pp. 557-562 ◽  
Author(s):  
Jean Pelletier ◽  
Bertrand Castro ◽  
Georges Roblot ◽  
Renée Wylde ◽  
Marie-Madeleine de Reviers
Keyword(s):  
Median Eminence ◽  
Binding Sites ◽  
Melatonin Receptors ◽  
Scatchard Analysis ◽  
Pars Tuberalis

Abstract. The present study was conducted to assess the binding of [125I]melatonin to frozen unfixed sections of pars tuberalis/median eminence tissue from Ile-de-France rams exposed or not exposed to light before slaughter. The specificity of [125I]melatonin binding to the pars tuberalis tissue was revealed by autoradiography and the magnitude of binding as related to the pars tuberalis area was determined after incubation and counting of pars tuberalis/median eminence sections. Subsequent studies with sections incubated with [125I]melatonin indicated that 1. the binding sites were saturable; 2. binding was stable for 24 h at 20°C, but unstable at 28 or 37°C; 3. melatonin and [12 7I]melatonin had a similar potency to compete with [125I]melatonin for binding sites, whereas other ligands such as serotonin or N-acetylserotonin were devoid of activity, and 4. by Scatchard analysis, the constant affinity Ka was found to be high in the 1010 l/mol range. Rams exposed to light throughout the night prior to slaughter presented a significant increase in the apparent number of [125I]melatonin binding sites in comparison to animals maintained under darkness (2.25±0.30 vs 1.01±0.17 fmol/mm2 pars tuberalis, p<0.01), whereas Ka values were similar in both groups. These results indicate the presence of true melatonin receptors in the pars tuberalis of the ram. Furthermore, they suggest that their apparent number is light-dependent.

scholarly journals Uncovering tissue-specific binding features from differential deep learning

2019 ◽  
Author(s):  
Mike Phuycharoen ◽  
Peyman Zarrineh ◽  
Laure Bridoux ◽  
Shilu Amin ◽  
Marta Losa ◽  
...  
Keyword(s):  
Deep Learning ◽  
Binding Sites ◽  
Specific Binding ◽  
Specific Expression ◽  
Tissue Specific ◽  
Dimensional Classification ◽  
Branchial Arches ◽  
Gradient Based ◽  
Differential Binding

ABSTRACTMotivationTranscription factors (TFs) can bind DNA in a cooperative manner, enabling a mutual increase in occupancy. Through this type of interaction, alternative binding sites can be preferentially bound in different tissues to regulate tissue-specific expression programmes. Recently, deep learning models have become state-of-the-art in various pattern analysis tasks, including applications in the field of genomics. We therefore investigate the application of convolutional neural network (CNN) models to the discovery of sequence features determining cooperative and differential TF binding across tissues.ResultsWe analyse ChIP-seq data from MEIS, TFs which are broadly expressed across mouse branchial arches, and HOXA2, which is expressed in the second and more posterior branchial arches. By developing models predictive of MEIS differential binding in all three tissues we are able to accurately predict HOXA2 co-binding sites. We evaluate transfer-like and multitask approaches to regularising the high-dimensional classification task with a larger regression dataset, allowing for creation of deeper and more accurate models. We test the performance of perturbation and gradient-based attribution methods in identifying the HOXA2 sites from differential MEIS data. Our results show that deep regularised models significantly outperform shallow CNNs as well as k-mer methods in the discovery of tissue-specific sites bound in vivo.AvailabilityFor implementation and models please visit https://doi.org/10.5281/zenodo.2635463.

Botulinum neurotoxin serotype D attacks neurons via two carbohydrate-binding sites in a ganglioside-dependent manner

2010 ◽  
Vol 431 (2) ◽  
pp. 207-216 ◽  
Author(s):  
Jasmin Strotmeier ◽  
Kwangkook Lee ◽  
Anne K. Völker ◽  
Stefan Mahrhold ◽  
Yinong Zong ◽  
...  
Keyword(s):  
Sialic Acid ◽  
Synaptic Vesicle ◽  
Motor Neurons ◽  
Binding Sites ◽  
Botulinum Neurotoxin ◽  
Specific Binding ◽  
Binding Pocket ◽  
The Other ◽  
Carbohydrate Binding ◽  
Dependent Manner

The extraordinarily high toxicity of botulinum neurotoxins primarily results from their specific binding and uptake into neurons. At motor neurons, the seven BoNT (botulinum neurotoxin) serotypes A–G inhibit acetylcholine release leading to flaccid paralysis. Uptake of BoNT/A, B, E, F and G requires a dual interaction with gangliosides and the synaptic vesicle proteins synaptotagmin or SV2 (synaptic vesicle glycoprotein 2), whereas little is known about the cell entry mechanisms of the serotypes C and D, which display the lowest amino acid sequence identity compared with the other five serotypes. In the present study we demonstrate that the neurotoxicity of BoNT/D depends on the presence of gangliosides by employing phrenic nerve hemidiaphragm preparations derived from mice expressing the gangliosides GM3, GM2, GM1 and GD1a, or only GM3 [a description of our use of ganglioside nomenclature is given in Svennerholm (1994) Prog. Brain Res. 101, XI–XIV]. High-resolution crystal structures of the 50 kDa cell-binding domain of BoNT/D alone and in complex with sialic acid, as well as biological analyses of single-site BoNT/D mutants identified two carbohydrate-binding sites. One site is located at a position previously identified in BoNT/A, B, E, F and G, but is lacking the conserved SXWY motif. The other site, co-ordinating one molecule of sialic acid, resembles the second ganglioside-binding pocket (the sialic-acid-binding site) of TeNT (tetanus neurotoxin).

Melatonin receptors are present in adult rat Leydig cells and are coupled through a pertussis toxin-sensitive G-protein

1997 ◽  
Vol 136 (6) ◽  
pp. 633-639 ◽  
Author(s):  
Sandra Valenti ◽  
Massimo Giusti ◽  
Roberta Guido ◽  
Giulio Giordano
Keyword(s):  
G Protein ◽  
Pertussis Toxin ◽  
Binding Sites ◽  
Leydig Cells ◽  
Melatonin Receptors ◽  
Scatchard Analysis ◽  
Adult Rat ◽  
Testosterone Secretion ◽  
High Affinity Binding Site ◽  
17 Hydroxyprogesterone

Abstract Previous studies have suggested that melatonin (MLT) acts directly on rat Leydig cells by modulating androgen production. In the present study, the site of action of MLT was investigated. The binding of 2-[125I]iodomelatonin (125I-MLT; 7–240 pmol/l) to Leydig cell membrane fragments was tested in the presence or absence of guanosine 5′-O-(3-thiotriphosphate) (GTP-γ-S; 50 μmol/l). Saturation studies and Scatchard analysis revealed the existence of a high-affinity binding site with a Bmax of 46·70± 3·50 fmol/mg protein and a Kd of 88·70±6·20 pmol/l; treatment with GTP-γ-S reduced the concentration of 125I-MLT binding sites (Bmax 34·03±4·50), while increasing the Kd to 106·5± 2·61 pmol/l. Pretreatment of the cells with pertussis toxin (PTX; 10 ng/ml for 16 h) resulted in a decreased binding of I-MLT and a lack of effect of GTP-γ-S. Moreover, the effect of MLT on testosterone secretion induced by LH (30 mIU/ml), forskolin (1 μmol/l) and LHRH (100 nmol/l) was studied after 3-h incubation of cells which had been precultured with or without PTX. The inhibition of testosterone secretion due to MLT administration was eliminated by PTX pretreatment during forskolin and LH, but not during LHRH administration. However, 17-hydroxyprogesterone levels were higher in all groups incubated in the presence of MLT, irrespective of PTX pretreatment. Our data suggest that: (a) MLT receptors are present on the membranes of adult rat Leydig cells; (b) they couple through PTX-sensitive G-protein-coupled binding sites; (c) the mechanism by which MLT blocks 17–20 desmolase enzymatic activity (thus leading to increased 17-hydroxyprogesterone levels), and testosterone secretion during LHRH stimulation is likely to depend on one or more different mechanism(s) of action. European Journal of Endocrinology 136 633–639

Daily Rhythms of Melatonin Binding Sites in the Rat Pars tuberalis and Suprachiasmatic Nuclei; Evidence for a Regulation of Melatonin Receptors by Melatonin Itself

1993 ◽  
Vol 57 (1) ◽  
pp. 120-126 ◽  
Author(s):  
François Gauer ◽  
Mireille Masson-Pévet ◽  
Debra Jean Skene ◽  
Berthe Vivien-Roels ◽  
Paul Pévet
Keyword(s):  
Binding Sites ◽  
Melatonin Receptors ◽  
Pars Tuberalis ◽  
Suprachiasmatic Nuclei ◽  
Daily Rhythms

Receptors for insulin-like growth factor II in the rat uterus: characterization and variation throughout the estrus cycle

1991 ◽  
Vol 124 (4) ◽  
pp. 434-441 ◽  
Author(s):  
Yallampalli Chandrasekhar ◽  
Satya Narayan ◽  
Pomila Singh ◽  
Manubai Nagamani
Keyword(s):  
Growth Factor ◽  
Binding Sites ◽  
Binding Capacity ◽  
Specific Binding ◽  
Cross Linking ◽  
Dependent Manner ◽  
Insulin Like Growth Factor ◽  
Estrus Cycle ◽  
Rat Uterus ◽  
Factor Ii

Abstract This study was undertaken to identify uterine insulin-like growth factor II receptors and examine the regulation of these receptor levels throughout the estrus cycle. We have demonstrated IGF-II receptors in crude uterine membranes by binding and cross-linking experiments. IGF-II binding to the rat uterine membranes displayed time and temperature dependence and maximum binding was achieved by 2 h at 22°C. Uterine IGF-II binding sites were specific for binding IGF-II peptide and demonstrated negligible binding affinity for IGF-I and no affinity for insulin. The specific anti-IGF-II receptor antibody, R-II-PAB1, blocked the specific [125I]IGF-II binding to uterine membranes in a dose-dependent manner. The characteristics of uterine IGF-II receptor are similar to those reported for other tissues, with a single class of high-affinity binding sites with an apparent dissociation constant of 1.2±0.5 nmol/l and βmax of 2.65±0.41 pmol/mg protein. Affinity cross-linking experiments indicated that the specific binding of [125I]IGF-II in the uterus is associated with a single band of protein with a mol wt of 250 kD. In mature cycling rats, the proestrus uterus had the lowest level of [125I]IGF-II binding per mg membrane protein, without changes in receptor affinity. However, because of greater yield of protein from proestrus uteri, the total [125I]IGF-II binding capacity of the uterus was similar to the other stages of the estrus cycle. These studies demonstrate the presence of authentic IGF-II receptors in the rat uterus and illustrate variations in the concentration of these receptors in the uterus throughout the estrus cycle.

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