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G B Ryan - One of the best experts on this subject based on the ideXlab platform.
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Podocyte architecture in Puromycin aminonucleoside-treated rats administered tungsten or allopurinol.
Experimental nephrology, 1995Co-Authors: Sharon D Ricardo, John F Bertram, G B RyanAbstract:The role of xanthine oxidase as a source of reactive oxygen species in Puromycin aminonucleoside nephrosis was examined. The effects of allopurinol (a xanthine oxidase inhibitor as well as a reactive oxygen species scavenging enzyme) and tungsten (a specific xanthine oxidase inhibitor) on glomerular epithelial cell ultrastructure, renal xanthine oxidase and xanthine dehydrogenase activity, and urinary protein excretion were examined in Puromycin aminonucleoside-treated rats. Co-administration of allopurinol to such rats reduced proteinuria by approximately 70% over the 10 days studied, and reduced the degree of glomerular epithelial cell foot process effacement at both 5 and 10 days, compared to rats that received Puromycin aminonucleoside alone. Unexpectedly, co-administration of allopurinol to Puromycin aminonucleoside-treated rats did not reduce xanthine oxidase activity; however, the combined activity of xanthine oxidase and xanthine dehydrogenase in such animals was reduced on day 5. Co-administration of tungsten to Puromycin aminonucleoside-treated rats did not reduce proteinuria or alter the number of filtration slits. Rats co-administered tungsten and Puromycin aminonucleoside had significantly reduced renal xanthine oxidase and combined xanthine oxidase and xanthine dehydrogenase activities on days 5 and 10, compared to rats treated with Puromycin aminonucleoside alone. These results provide evidence that the protection provided by allopurinol in Puromycin aminonucleoside-treated rats is due to the antioxidant properties of allopurinol, rather than to its activities as a xanthine oxidase inhibitor.
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antioxidants protect podocyte foot processes in Puromycin aminonucleoside treated rats
Journal of The American Society of Nephrology, 1994Co-Authors: Sharon D Ricardo, John F Bertram, G B RyanAbstract:Whether a reduction in urinary protein excretion in rats coadministered Puromycin aminonucleoside and antioxidants was associated with a reduction in alterations to glomerular epithelial cell (podocyte) ultrastructure was examined. Daily urinary protein excretion was measured in rats that received a single i.v. injection of saline or Puromycin aminonucleoside with or without coadministration of antioxidants. The coadministration of alpha-tocopherol/ascorbic acid, dimethyl thiourea, or superoxide dismutase to Puromycin aminonucleoside-treated rats reduced proteinuria by approximately 90, 40, and 60%, respectively, over the 18-day period studied. For a second group of rats, daily urinary protein excretion was measured and kidneys were processed for light microscopy and transmission and scanning electron microscopy 4, 5, and 10 days after injection. Transmission electron microscopic morphometric analysis of glomeruli from Puromycin aminonucleoside-treated rats coadministered antioxidants revealed significantly reduced foot process effacement on Days, 4, 5, and 10 compared with rats that received Puromycin aminonucleoside alone. Thus, at Day 10, Puromycin aminonucleoside-treated rats coadministered alpha-tocopherol/ascorbic acid, dimethyl thiourea, or superoxide dismutase contained 90, 74, and 88% (P
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Antioxidants protect podocyte foot processes in Puromycin aminonucleoside-treated rats.
Journal of the American Society of Nephrology : JASN, 1994Co-Authors: Sharon D Ricardo, John F Bertram, G B RyanAbstract:Whether a reduction in urinary protein excretion in rats coadministered Puromycin aminonucleoside and antioxidants was associated with a reduction in alterations to glomerular epithelial cell (podocyte) ultrastructure was examined. Daily urinary protein excretion was measured in rats that received a single i.v. injection of saline or Puromycin aminonucleoside with or without coadministration of antioxidants. The coadministration of alpha-tocopherol/ascorbic acid, dimethyl thiourea, or superoxide dismutase to Puromycin aminonucleoside-treated rats reduced proteinuria by approximately 90, 40, and 60%, respectively, over the 18-day period studied. For a second group of rats, daily urinary protein excretion was measured and kidneys were processed for light microscopy and transmission and scanning electron microscopy 4, 5, and 10 days after injection. Transmission electron microscopic morphometric analysis of glomeruli from Puromycin aminonucleoside-treated rats coadministered antioxidants revealed significantly reduced foot process effacement on Days, 4, 5, and 10 compared with rats that received Puromycin aminonucleoside alone. Thus, at Day 10, Puromycin aminonucleoside-treated rats coadministered alpha-tocopherol/ascorbic acid, dimethyl thiourea, or superoxide dismutase contained 90, 74, and 88% (P < 0.01 in all cases) more glomerular epithelial cell filtration slits per unit length of glomerular basement membrane than rats treated with Puromycin aminonucleoside alone. In contrast, by scanning electron microscopy, the antioxidants were found to provide no protection against the changes occurring in glomerular epithelial cell bodies and major processes. These results provide further evidence of a role for reactive oxygen species in Puromycin aminonucleoside nephrosis and indicate that the antioxidants provide protection against the changes occurring in glomerular epithelial cell foot processes.
Denis Drainas - One of the best experts on this subject based on the ideXlab platform.
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kinetics of inhibition of ribonuclease p activity by peptidyltransferase inhibitors effect of antibiotics on rnase p
Molecular Biology Reports, 2003Co-Authors: Dimitra Kalavrizioti, Denis Drainas, Constantinos Stathopoulos, Antigoni Tsagla, Apostolos Tekos, Anastassios VourekasAbstract:A cell-free system derived from Dictyostelium discoideum has been used to study the kinetics of inhibition of RNase P by Puromycin, amicetin and blasticidin S. Detailed kinetic analysis showed that the type of inhibition of RNase P activity by Puromycin is simple competitive, whereas the type of inhibition by amicetin and blasticidin S is simple non-competitive. On the basis of Ki values amicetin is stronger inhibitor than Puromycin and blasticidin S.
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kinetics of inhibition of ribonuclease p activity by peptidyltransferase inhibitors
2003Co-Authors: Dimitra Kalavrizioti, Apostolos Tekos, Antigoni Tsagla, Anastassios Vourekas, Denis DrainasAbstract:A cell-free system derived from Dictyostelium discoideum has been used to study the kinetics of inhibition of RNase P by Puromycin, amicetin and blasticidin S. Detailed kinetic analysis showed that the type of inhibition of RNase P activity by Puromycin is simple competitive, whereas the type of inhibition by amicetin and blasticidin S is simple non-competitive. On the basis of Ki values amicetin is stronger inhibitor than Puromycin and blasticidin S.
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Effect of peptidyltransferase inhibitors on ribonuclease P activity from Dictyostelium discoideum
2000Co-Authors: Constantinos Stathopoulos, Antigoni Tsagla, Apostolos Tekos, Denis DrainasAbstract:The effect of several peptidyltransferase inhibitors on ribonuclease P activity from Dictyostelium discoideum was investigated. Among the inhibitors tested Puromycin, amicetin and blasticidin S revealed a dose-dependent inhibition of tRNA maturation. Blasticidin S and amicetin do not compete with Puromycin for the same site on the enzyme, suggesting the existence of distinct antibiotic binding sites on D. discoideum RNase P. Inhibition experiments further indicate that binding sites for blasticidin S and amicetin overlap.
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Effect of peptidyltransferase inhibitors on ribonuclease P activity from Dictyostelium discoideum. Effect of antibiotics on RNase P.
Molecular biology reports, 2000Co-Authors: Constantinos Stathopoulos, Antigoni Tsagla, Apostolos Tekos, Denis DrainasAbstract:The effect of several peptidyltransferase inhibitors on ribonuclease P activity from Dictyostelium discoideum was investigated. Among the inhibitors tested Puromycin, amicetin and blasticidin S revealed a dose-dependent inhibition of tRNA maturation. Blasticidin S and amicetin do not compete with Puromycin for the same site on the enzyme, suggesting the existence of distinct antibiotic binding sites on D. discoideum RNase P. Inhibition experiments further indicate that binding sites for blasticidin S and amicetin overlap.
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bimodal action of spermine on ribosomal peptidyltransferase at low concentration of magnesium ions
Biochimica et Biophysica Acta, 1994Co-Authors: Denis Drainas, Dimitrios L KalpaxisAbstract:At 6 mM Mg2+, submillimolar concentrations of spermine affect the end-point as well as the kinetic phase of Puromycin reaction in a cell-free system from Escherichia coli. When the ternary complex AcPhe-tRNA-poly(U)-ribosome (complex C) is formed in the absence of ribosomal wash (FWR fraction), the final degree of AcPhe-Puromycin synthesis is raised from 12% to 60%, as the concentration of spermine increases from zero to 200 microM. However, spermine displays partial noncompetitive inhibition at the kinetic phase of the reaction. The inhibitory effect of spermine is related with its binding to AcPhe-tRNA. When complex C is formed in the presence of FWR fraction, spermine slightly affects the final degree of Puromycin synthesis is markedly stimulated by the addition of relatively low concentrations of spermine. Kinetic analysis of the activation phase revealed that spermine attached on a specific site of complex C, acts as a nonessential, partial noncompetitive activator. The stimulatory effect of spermine seems to be due to its interaction with ribosomes. Further additions of spermine cause partial noncompetitive inhibition on the Puromycin reaction. This result suggests that complex C possesses a second binding site, responsible for the inhibitory effect of spermine. Both activator and inhibitor sites can be occupied by spermine at the same time.
Sharon D Ricardo - One of the best experts on this subject based on the ideXlab platform.
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Podocyte architecture in Puromycin aminonucleoside-treated rats administered tungsten or allopurinol.
Experimental nephrology, 1995Co-Authors: Sharon D Ricardo, John F Bertram, G B RyanAbstract:The role of xanthine oxidase as a source of reactive oxygen species in Puromycin aminonucleoside nephrosis was examined. The effects of allopurinol (a xanthine oxidase inhibitor as well as a reactive oxygen species scavenging enzyme) and tungsten (a specific xanthine oxidase inhibitor) on glomerular epithelial cell ultrastructure, renal xanthine oxidase and xanthine dehydrogenase activity, and urinary protein excretion were examined in Puromycin aminonucleoside-treated rats. Co-administration of allopurinol to such rats reduced proteinuria by approximately 70% over the 10 days studied, and reduced the degree of glomerular epithelial cell foot process effacement at both 5 and 10 days, compared to rats that received Puromycin aminonucleoside alone. Unexpectedly, co-administration of allopurinol to Puromycin aminonucleoside-treated rats did not reduce xanthine oxidase activity; however, the combined activity of xanthine oxidase and xanthine dehydrogenase in such animals was reduced on day 5. Co-administration of tungsten to Puromycin aminonucleoside-treated rats did not reduce proteinuria or alter the number of filtration slits. Rats co-administered tungsten and Puromycin aminonucleoside had significantly reduced renal xanthine oxidase and combined xanthine oxidase and xanthine dehydrogenase activities on days 5 and 10, compared to rats treated with Puromycin aminonucleoside alone. These results provide evidence that the protection provided by allopurinol in Puromycin aminonucleoside-treated rats is due to the antioxidant properties of allopurinol, rather than to its activities as a xanthine oxidase inhibitor.
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antioxidants protect podocyte foot processes in Puromycin aminonucleoside treated rats
Journal of The American Society of Nephrology, 1994Co-Authors: Sharon D Ricardo, John F Bertram, G B RyanAbstract:Whether a reduction in urinary protein excretion in rats coadministered Puromycin aminonucleoside and antioxidants was associated with a reduction in alterations to glomerular epithelial cell (podocyte) ultrastructure was examined. Daily urinary protein excretion was measured in rats that received a single i.v. injection of saline or Puromycin aminonucleoside with or without coadministration of antioxidants. The coadministration of alpha-tocopherol/ascorbic acid, dimethyl thiourea, or superoxide dismutase to Puromycin aminonucleoside-treated rats reduced proteinuria by approximately 90, 40, and 60%, respectively, over the 18-day period studied. For a second group of rats, daily urinary protein excretion was measured and kidneys were processed for light microscopy and transmission and scanning electron microscopy 4, 5, and 10 days after injection. Transmission electron microscopic morphometric analysis of glomeruli from Puromycin aminonucleoside-treated rats coadministered antioxidants revealed significantly reduced foot process effacement on Days, 4, 5, and 10 compared with rats that received Puromycin aminonucleoside alone. Thus, at Day 10, Puromycin aminonucleoside-treated rats coadministered alpha-tocopherol/ascorbic acid, dimethyl thiourea, or superoxide dismutase contained 90, 74, and 88% (P
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Antioxidants protect podocyte foot processes in Puromycin aminonucleoside-treated rats.
Journal of the American Society of Nephrology : JASN, 1994Co-Authors: Sharon D Ricardo, John F Bertram, G B RyanAbstract:Whether a reduction in urinary protein excretion in rats coadministered Puromycin aminonucleoside and antioxidants was associated with a reduction in alterations to glomerular epithelial cell (podocyte) ultrastructure was examined. Daily urinary protein excretion was measured in rats that received a single i.v. injection of saline or Puromycin aminonucleoside with or without coadministration of antioxidants. The coadministration of alpha-tocopherol/ascorbic acid, dimethyl thiourea, or superoxide dismutase to Puromycin aminonucleoside-treated rats reduced proteinuria by approximately 90, 40, and 60%, respectively, over the 18-day period studied. For a second group of rats, daily urinary protein excretion was measured and kidneys were processed for light microscopy and transmission and scanning electron microscopy 4, 5, and 10 days after injection. Transmission electron microscopic morphometric analysis of glomeruli from Puromycin aminonucleoside-treated rats coadministered antioxidants revealed significantly reduced foot process effacement on Days, 4, 5, and 10 compared with rats that received Puromycin aminonucleoside alone. Thus, at Day 10, Puromycin aminonucleoside-treated rats coadministered alpha-tocopherol/ascorbic acid, dimethyl thiourea, or superoxide dismutase contained 90, 74, and 88% (P < 0.01 in all cases) more glomerular epithelial cell filtration slits per unit length of glomerular basement membrane than rats treated with Puromycin aminonucleoside alone. In contrast, by scanning electron microscopy, the antioxidants were found to provide no protection against the changes occurring in glomerular epithelial cell bodies and major processes. These results provide further evidence of a role for reactive oxygen species in Puromycin aminonucleoside nephrosis and indicate that the antioxidants provide protection against the changes occurring in glomerular epithelial cell foot processes.
Peter A. Sims - One of the best experts on this subject based on the ideXlab platform.
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Elongation inhibitors do not prevent the release of puromycylated nascent polypeptide chains from ribosomes.
eLife, 2020Co-Authors: Benjamin D. Hobson, Linghao Kong, Erik W. Hartwick, Ruben L. Gonzalez, Peter A. SimsAbstract:Puromycin is an amino-acyl transfer RNA analog widely employed in studies of protein synthesis. Since Puromycin is covalently incorporated into nascent polypeptide chains, anti-Puromycin immunofluorescence enables visualization of nascent protein synthesis. A common assumption in studies of local messenger RNA translation is that the anti-Puromycin staining of puromycylated nascent polypeptides in fixed cells accurately reports on their original site of translation, particularly when ribosomes are stalled with elongation inhibitors prior to Puromycin treatment. However, when we attempted to implement a proximity ligation assay to detect ribosome-Puromycin complexes, we found no evidence to support this assumption. We further demonstrated, using biochemical assays and live cell imaging of nascent polypeptides in mammalian cells, that puromycylated nascent polypeptides rapidly dissociate from ribosomes even in the presence of elongation inhibitors. Our results suggest that attempts to define precise subcellular translation sites using anti-Puromycin immunostaining may be confounded by release of puromycylated nascent polypeptide chains prior to fixation.
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Elongation Inhibitors do not Prevent the Release of Puromycylated Nascent Polypeptide Chains from Ribosomes
2020Co-Authors: Benjamin D. Hobson, Linghao Kong, Erik W. Hartwick, Ruben L. Gonzalez, Peter A. SimsAbstract:Puromycin is an amino-acyl transfer RNA analog widely employed in studies of protein synthesis. Since Puromycin is covalently incorporated into nascent polypeptide chains, anti-Puromycin immunofluorescence enables visualization of nascent protein synthesis. A common assumption in studies of local messenger RNA translation is that the anti-Puromycin staining of puromycylated nascent polypeptides in fixed cells accurately reports on their original site of translation, particularly when ribosomes are stalled with elongation inhibitors prior to Puromycin treatment. However, when we attempted to implement a proximity ligation assay to detect ribosome-Puromycin complexes, we found no evidence to support this assumption. We further demonstrated, using biochemical assays and live cell imaging of nascent polypeptides, that puromycylated nascent polypeptides rapidly dissociate from ribosomes even in the presence of elongation inhibitors. Our results suggest that attempts to define precise subcellular translation sites using anti-Puromycin immunostaining may be confounded by release of puromycylated nascent polypeptide chains prior to fixation.
John F Bertram - One of the best experts on this subject based on the ideXlab platform.
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Podocyte architecture in Puromycin aminonucleoside-treated rats administered tungsten or allopurinol.
Experimental nephrology, 1995Co-Authors: Sharon D Ricardo, John F Bertram, G B RyanAbstract:The role of xanthine oxidase as a source of reactive oxygen species in Puromycin aminonucleoside nephrosis was examined. The effects of allopurinol (a xanthine oxidase inhibitor as well as a reactive oxygen species scavenging enzyme) and tungsten (a specific xanthine oxidase inhibitor) on glomerular epithelial cell ultrastructure, renal xanthine oxidase and xanthine dehydrogenase activity, and urinary protein excretion were examined in Puromycin aminonucleoside-treated rats. Co-administration of allopurinol to such rats reduced proteinuria by approximately 70% over the 10 days studied, and reduced the degree of glomerular epithelial cell foot process effacement at both 5 and 10 days, compared to rats that received Puromycin aminonucleoside alone. Unexpectedly, co-administration of allopurinol to Puromycin aminonucleoside-treated rats did not reduce xanthine oxidase activity; however, the combined activity of xanthine oxidase and xanthine dehydrogenase in such animals was reduced on day 5. Co-administration of tungsten to Puromycin aminonucleoside-treated rats did not reduce proteinuria or alter the number of filtration slits. Rats co-administered tungsten and Puromycin aminonucleoside had significantly reduced renal xanthine oxidase and combined xanthine oxidase and xanthine dehydrogenase activities on days 5 and 10, compared to rats treated with Puromycin aminonucleoside alone. These results provide evidence that the protection provided by allopurinol in Puromycin aminonucleoside-treated rats is due to the antioxidant properties of allopurinol, rather than to its activities as a xanthine oxidase inhibitor.
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antioxidants protect podocyte foot processes in Puromycin aminonucleoside treated rats
Journal of The American Society of Nephrology, 1994Co-Authors: Sharon D Ricardo, John F Bertram, G B RyanAbstract:Whether a reduction in urinary protein excretion in rats coadministered Puromycin aminonucleoside and antioxidants was associated with a reduction in alterations to glomerular epithelial cell (podocyte) ultrastructure was examined. Daily urinary protein excretion was measured in rats that received a single i.v. injection of saline or Puromycin aminonucleoside with or without coadministration of antioxidants. The coadministration of alpha-tocopherol/ascorbic acid, dimethyl thiourea, or superoxide dismutase to Puromycin aminonucleoside-treated rats reduced proteinuria by approximately 90, 40, and 60%, respectively, over the 18-day period studied. For a second group of rats, daily urinary protein excretion was measured and kidneys were processed for light microscopy and transmission and scanning electron microscopy 4, 5, and 10 days after injection. Transmission electron microscopic morphometric analysis of glomeruli from Puromycin aminonucleoside-treated rats coadministered antioxidants revealed significantly reduced foot process effacement on Days, 4, 5, and 10 compared with rats that received Puromycin aminonucleoside alone. Thus, at Day 10, Puromycin aminonucleoside-treated rats coadministered alpha-tocopherol/ascorbic acid, dimethyl thiourea, or superoxide dismutase contained 90, 74, and 88% (P
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Antioxidants protect podocyte foot processes in Puromycin aminonucleoside-treated rats.
Journal of the American Society of Nephrology : JASN, 1994Co-Authors: Sharon D Ricardo, John F Bertram, G B RyanAbstract:Whether a reduction in urinary protein excretion in rats coadministered Puromycin aminonucleoside and antioxidants was associated with a reduction in alterations to glomerular epithelial cell (podocyte) ultrastructure was examined. Daily urinary protein excretion was measured in rats that received a single i.v. injection of saline or Puromycin aminonucleoside with or without coadministration of antioxidants. The coadministration of alpha-tocopherol/ascorbic acid, dimethyl thiourea, or superoxide dismutase to Puromycin aminonucleoside-treated rats reduced proteinuria by approximately 90, 40, and 60%, respectively, over the 18-day period studied. For a second group of rats, daily urinary protein excretion was measured and kidneys were processed for light microscopy and transmission and scanning electron microscopy 4, 5, and 10 days after injection. Transmission electron microscopic morphometric analysis of glomeruli from Puromycin aminonucleoside-treated rats coadministered antioxidants revealed significantly reduced foot process effacement on Days, 4, 5, and 10 compared with rats that received Puromycin aminonucleoside alone. Thus, at Day 10, Puromycin aminonucleoside-treated rats coadministered alpha-tocopherol/ascorbic acid, dimethyl thiourea, or superoxide dismutase contained 90, 74, and 88% (P < 0.01 in all cases) more glomerular epithelial cell filtration slits per unit length of glomerular basement membrane than rats treated with Puromycin aminonucleoside alone. In contrast, by scanning electron microscopy, the antioxidants were found to provide no protection against the changes occurring in glomerular epithelial cell bodies and major processes. These results provide further evidence of a role for reactive oxygen species in Puromycin aminonucleoside nephrosis and indicate that the antioxidants provide protection against the changes occurring in glomerular epithelial cell foot processes.
