Localization and properties of bovine photoreceptor 5'-nucleotidase

Yu, Longbo; Fager, Roger S.

doi:10.1152/ajpcell.1983.244.1.c89

Cited by 74 publications

(4 citation statements)

References 29 publications

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“…In the outer segment of vertebral photoreceptor cells, the unique properties of 5 Ј-NT have been investigated biochemically. Both membrane-bound and soluble 5 Ј-NTs are present Shichi 1981,1982;Yu and Fager 1983). From biochemical studies, the catalytic site of the soluble 5 Ј-NT is on the extradiscal side of the disk membrane whereas that of the membrane-bound enzyme is on the intradiscal side Shichi 1981,1982;Yu and Fager 1983).…”

Section: Discussionmentioning

confidence: 99%

“…Both membrane-bound and soluble 5 Ј-NTs are present Shichi 1981,1982;Yu and Fager 1983). From biochemical studies, the catalytic site of the soluble 5 Ј-NT is on the extradiscal side of the disk membrane whereas that of the membrane-bound enzyme is on the intradiscal side Shichi 1981,1982;Yu and Fager 1983). The soluble form is believed to be of particular functional importance for the phototransduction system of living photoreceptor cells, whereas it accounts for about 25% of the total 5 Ј-NT activity (Fukui and Shichi 1982).…”

Section: Discussionmentioning

confidence: 99%

“…The soluble form is believed to be of particular functional importance for the phototransduction system of living photoreceptor cells, whereas it accounts for about 25% of the total 5 Ј-NT activity (Fukui and Shichi 1982). On the other hand, the membrane-bound 5 Ј-NT is considered a nonfunctional enzyme in situ (Yu and Fager 1983). Although the biochemical topology of 5 Ј-NT in the outer segment has been revealed, the cytochemical topology of 5 Ј-NT is controversial.…”

Section: Discussionmentioning

confidence: 99%

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5′-Nucleotidase in Rat Photoreceptor Cells and Pigment Epithelial Cells Processed by Rapid-freezing Enzyme Cytochemistry

Tajima

1998

J Histochem Cytochem.

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SUMMARYThis report describes the subcellular distribution of 5 Ј-nucleotidase (5 Ј-NT) in rat photoreceptor cells and pigment epithelial cells processed by rapid-freeze enzyme cytochemistry. There was a striking difference in the ultrastructural localization of 5 Ј-NT activity between rod outer segments after freeze-substitution fixation and conventional fixation. By rapid-freezing enzyme cytochemistry, 5 Ј-NT activity was localized in the extradiscal space of intact nonvacuolated discs, whereas by conventional cytochemistry it was shown in the intradiscal space of artifactual vacuolated discs. In the freeze-substituted retinal cells, an appreciable difference in functional 5 Ј-NT molecules was also found. The soluble 5 Ј-NT on the cytoplasmic side of the disc membrane was vital in the rod outer segments, whereas the membrane-bound ecto-5 Ј-NT on the exoplasmic (external) surface of the apical process was active in the pigment epithelial cells. Rapid-freezing enzyme cytochemistry should be worth employing as a method to reveal the fine localization of enzyme activity at the level of cell ultrastructures, which are poorly preserved by conventional fixation, and should provide information approximate to that in living cells. Electron microscopic enzyme cytochemistry is a morphological technique for analyzing topology and dynamics of enzymes in situ. Although chemical fixation is always essential in this technique, immobilization of cell ultrastructures containing enzyme molecules by conventional fixation often reveals contradictory preservation of enzyme activity (i.e., inactivation of enzyme activity vs artifactual diffusion of enzyme activity). Freeze-substitution fixation is a technique that overcomes the obstacles of conventional fixation. Rapid-freezing enzyme cytochemistry, which combines rapid-freezing, freeze-substitution fixation, and subsequent enzyme cytochemistry, enables us to reveal the precise localization of enzymes on well-preserved cell structures (Klaushofer and von Mayersbach 1979; Saito 1989,1992;Robinson and Karnovsky 1991;Saito and Takizawa 1991).In retinal photoreceptor cells, the enzyme 5 Ј-nucleotidase (5 Ј-NT) is one of the enzymes in the cGMP metabolism cascade of the vertebrate visual transduction system. Although some researchers have studied the enzyme cytochemical localization of 5 Ј-NT in retinal rod cells processed by conventional fixation (Kreutzberg and Hussain 1984;Hussain and Baydoun 1985;Saito 1991), they have not been in good agreement on its localization. One of the reasons is that rod cells are one of the most delicate cells to treat by conventional fixation and morphological modifications during fixation are often encountered. Yamada (1982) showed the fine structure of intact discs in the rod outer segment processed by rapid-freezing and subsequent freezesubstitution fixation. Therefore, it would be interesting to detect 5 Ј-NT activity in freeze-substituted photoreceptor cells. This article presents the ultrastructural localization of 5 Ј-NT in rat photoreceptor cells an...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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5′-Nucleotidase in Rat Photoreceptor Cells and Pigment Epithelial Cells Processed by Rapid-freezing Enzyme Cytochemistry

Tajima

1998

J Histochem Cytochem.

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“…There are some problems with the 5'-nucleotidase in this context: its location in ROS (cytoplasmicexposed or not, or mixed?) is still controversial [7, 8, 241, and 5'-nucleotidase is strongly inhibited by ATP, having an inhibitory constant as low as 18 pM ATP [8]. Moreover, it would seem more useful for the rod cell to have its guanosine nucleotides recycled rather than degraded.…”

Section: Discussionmentioning

confidence: 99%

Purification and properties of guanylate kinase from bovine retinas and rod outer segments

Hall¹,

Kühn²

1986

European Journal of Biochemistry

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The presence of three soluble nucleotide phosphotransferases in bovine rod outer segments was demonstrated: guanylate kinase (EC 2.7.4.8), nucleoside-diphosphate kinase (EC 2.7.4.6) and adenylate kinase (EC 2.7.4.3).The enzyme guanylate kinase, which catalyzes the reaction GMP + ATP GDP + ADP, was purified to homogeneity from isolated bovine rod outer segments as well as from bovine retinas. The enzyme preparations obtained from both sources are identical in their chromatographic properties, molecular mass (20-23 kDa for both native enzyme and dodecylsulfate-denatured polypeptide), K , values (13 pM for GMP and 430 pM for ATP), specific activities, and nucleotide specificities. The enzyme's turnover number was estimated to be 130 s-'. The minimum amount of enzyme found in rod outer segments is about 1 copy per 800 rhodopsin molecules. The role of the enzyme in the cyclic GMP cycle in rod outer segments is discussed.Cyclic GMP has long been known to play an important role in rod outer segments (ROS). Early studies showed that levels of cGMP are regulated by light [l, 21 and recent studies have now implicated cGMP as the internal messenger in ROS [3,4]. The metabolism of cGMP in the rod cell, then, becomes a very important question. It is known that absorption of light by rhodopsin leads to the activation of a GTP-binding protein (transducin), which in turn activates a phosphodiesterase which hydrolyzes cGMP to 5'-GMP [5]. Synthesis of cGMP is accomplished from GTP by the membrane-bound enzyme guanylate cyclase [6]. 5'-Nucleotidase has also been characterized in ROS [7, 81; this enzyme hydrolyzes 5'-GMP to guanosine. Less is known about the two anabolistic enzymes, guanylate kinase (5'-GMP + ATP + GDP + ADP) and nucleoside-diphosphate kinase (NDP + ATP e NTP + ADP). The activities of these two enzymes have been measured in frozen retinal layers [9], and in preparations of truncated ROS [lo], but the enzymes themselves have not been further characterized. Evidence for the transfer of highenergy phosphate between nucleotides has also been obtained from studies on the recycling of NADPH in rod outer segments [loa]. We report here on the purification and properties of guanylate kinase, the enzyme that specifically

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Distribution of mesothelial stomata in the rat

et al. 1994

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In 15 rats the morphology and distribution of stomata and mesothelial cells of the diaphragm and of different serosal areas were studied by light and electron microscopy. Cell surface patterns, identified by SEM, vaned according to location. Mesothelial cells of the diaphragmatic peritoneum were either cuboidal or flattened. Stomata were found only among cuboidal cells and were consistently and exclusively observed on the muscular part of the diaphragm, predominantly on the peritoneal surface. Stomata appeared oval or rounded, 4-10 bm in diameter. When India ink was introduced into the peritoneal cavity as a tracer, carbon particles were rapidly removed through stomata into the subjacent lymphatic lacunae of the diaphragm. Mesothelium overlying the diaphragm differed morphologically from that over the remaining peritoneal and pleural cavities. Mesothelium of the latter formed a continuous sheet of flattened cells whose surfaces were studded with numerous microvilli. Stomata were absent. It is suggested that stomata are exclusive to the diaphragm and may serve as the main drainage channels for fluid and particles from the peritoneal cavity. This has important clinical implications. Thus stomata could represent a liability by providing a route for the escape of tumor cells and pathogens from the peritoneal cavity. However, they may represent an asset by providing an accessible route for blood transfusion and for peritoneal dialysis in treating chronic renal failure.

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Localization and properties of bovine photoreceptor 5'-nucleotidase

Cited by 74 publications

References 29 publications

5′-Nucleotidase in Rat Photoreceptor Cells and Pigment Epithelial Cells Processed by Rapid-freezing Enzyme Cytochemistry

5′-Nucleotidase in Rat Photoreceptor Cells and Pigment Epithelial Cells Processed by Rapid-freezing Enzyme Cytochemistry

Purification and properties of guanylate kinase from bovine retinas and rod outer segments

Distribution of mesothelial stomata in the rat

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