Inhibition of nuclear-protein phosphorylation in vitro by beryllium

1983 ◽  
Vol 3 (10) ◽  
pp. 955-962 ◽  
Author(s):  
Betsan E. Williams ◽  
David N. Skilleter
Keyword(s):  
Protein Phosphorylation ◽  
Rat Liver ◽  
Cyclic Nucleotide ◽  
Binding Sites ◽  
Nuclear Protein ◽  
Adult Rat ◽  
Liver Nuclei ◽  
Protein Dephosphorylation ◽  
Independent Protein

Endogenous cyclic-nucleotide-independent protein phosphorylation by ATP at pH 6.5 in adult rat liver nuclei in vitro is inhibited by beryllium (Be2+), but under the same conditions nuclear-protein dephosphorylation appears to be insensitive to Be2+. Prior incubation of nuclei with Be2+ is necessary to demonstrate the inhibition of phosphorylation, which increases as the pH is decreased from pH 8.0 to 6.5. The extent of inhibition can be related to the level of nuclear Be2+ binding and, evidence suggests, may be caused by direct or indirect interference by Be2+ with Mg2+ binding sites normally required to facilitate protein phosphorylation.

scholarly journals The association in vitro of polyribosomes with ribonuclease-treated derivatives of hepatic rough endoplasmic reticulum. Characteristics of the membrane binding sites and factors influencing association

1971 ◽  
Vol 125 (1) ◽  
pp. 67-79 ◽  
Author(s):  
T. K. Shires ◽  
L. Narurkar ◽  
H. C. Pitot
Keyword(s):  
Rat Liver ◽  
Binding Sites ◽  
Binding Capacity ◽  
Trypsin Treatment ◽  
Partial Removal ◽  
Adult Rat ◽  
Microsomal Membranes ◽  
Derivatives Of ◽  
Membrane Binding Sites

1. Pancreatic ribonuclease in dilute EDTA has been shown to condition rough-microsomal membranes from adult rat liver to accept exogenously added rat liver polyribosomes in vitro at 0–4°C. Treated smooth membranes would not significantly interact with polyribosomes. 2. The conditioning process decreased the membrane RNA content and removed polyribosomes from vesicle surfaces as viewed electron-microscopically. 3. Binding to these conditioned membranes was shown to be uninfluenced by changes of temperature (0–37°C) and pH (6.9–7.8) or the presence of cell sap, but was inhibited by increasing the concentration of potassium chloride. 4. Possession of a polyribosome-binding capacity by conditioned rough membranes was not dependent on adventitious materials that could be dislodged by high ionic strengths. 5. Trypsin treatment under mild conditions destroyed the binding capacity of ribonuclease-conditioned rough membranes. 6. A 2–10S residual RNA was recovered from ribonuclease-conditioned membranes, but its partial removal had no effect on the capacity of membranes to accept polyribosomes. However, some role for this residual RNA in attaching polyribosomes could not be discounted. 7. Evidence is considered that polyribosome-binding sites are intrinsic features of conditioned membranes isolated from rough-microsomal fractions, and that long-range ionic bonding is a primary factor in polyribosome interaction with these binding sites.

scholarly journals Inhibition of in vitro nuclear transport by a lectin that binds to nuclear pores.

1987 ◽  
Vol 104 (2) ◽  
pp. 189-200 ◽  
Author(s):  
D R Finlay ◽  
D D Newmeyer ◽  
T M Price ◽  
D J Forbes
Keyword(s):  
Rat Liver ◽  
Protein Transport ◽  
Nuclear Protein ◽  
Nuclear Transport ◽  
Nuclear Pore ◽  
Major Mechanism ◽  
Rat Liver Nuclei ◽  
Egg Extract ◽  
Liver Nuclei

Selective transport of proteins is a major mechanism by which biochemical differences are maintained between the cytoplasm and nucleus. To begin to investigate the molecular mechanism of nuclear transport, we used an in vitro transport system composed of a Xenopus egg extract, rat liver nuclei, and a fluorescently labeled nuclear protein, nucleoplasmin. With this system, we screened for inhibitors of transport. We found that the lectin, wheat germ agglutinin (WGA), completely inhibits the nuclear transport of fluorescently labeled nucleoplasmin. No other lectin tested affected nuclear transport. The inhibition by WGA was not seen when N-acetylglucosamine was present and was reversible by subsequent addition of sugar. When rat liver nuclei that had been incubated with ferritin-labeled WGA were examined by electron microscopy, multiple molecules of WGA were found bound to the cytoplasmic face of each nuclear pore. Gel electrophoresis and nitrocellulose transfer identified one major and several minor nuclear protein bands as binding 125I-labeled WGA. The most abundant protein of these, a 63-65-kD glycoprotein, is a candidate for the inhibitory site of action of WGA on nuclear protein transport. WGA is the first identified inhibitor of nuclear protein transport and interacts directly with the nuclear pore.

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