In quail oviducts the rate of synthesis of avidin, the biological end-point marker for the molecular events caused by progesterone, decreases with age. The cause of the reduced capacity of avidin induction has been studied on the molecular biological level polyadenylation of RNA which is one step in the process of post-transcriptional modification of heterogeneous nuclear RNA resulting in the formation of functional mRNA molecules. This novel approach was biochemically possible after the discovery of the poly(A) anabolic enzyme (poly(A) polymerase) and the two poly(A) catabolic enzymes (endoribonuclease IV and 5'-exoribonuclease). These enzymes are involved in the synthesis and degradation of the poly(A) segment of mRNA in vitro and most likely also in poly(A) metabolism in intact cell systems. Enzymatically controlled poly(A) metabolism of mRNA is regulated by the following interrelations: poly(A)-associated proteins and endoribonuclease IV; labilizing factor and poly(A)-associated proteins; 5'-exoribonuclease in cooperation with endoribonuclease IV and poly(A) polymerase. A close correlation between high levels of poly(A) catabolic enzymes and low rate of protein synthesis which was established in cell culture systems, seems also to be partially the biochemical cause for the reduced avidin synthesis in aging quail oviduct.

Aging↗

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Age-dependent gene induction in quail oviduct. XI. Alterations on the post-transcriptional level (analytical aspect).

For the elucidation of the age-dependent reduction of avidin synthesis in the oviduct of quails studies on on the level of post-transcriptional modification of mRNA were performed. Incorporation of [3H] adenosine into mRNA is drastically reduced in senescent animals compared to adult animals. Double labeling of mRNA with [3H]-adenosine and [3H] uridine revealed that in the presence of actinomycin D the incorporation ratio adenosine/uridine into mRNA increases to higher values in the case of adult animals compared with the ratio determined for mRNA from senescent animals. These data support the assumption that in senescent animals the length of the poly(A) sequence on mRNA might be shorter. Because of the low efficiency of label incorporation into poly(A) in whole animals, a method for directly introducing a radioactive label in vitro into the isolated poly(A) was developed for the estimation of the length of the poly(A) segments. A randomly labeled product with no apparent degradation is achieved by methylation of poly(A) with [3H] dimethyl sulphate. Therefore the method is a powerful tool for the detection and size determination of small amounts of poly(A). Size determination of poly(A) from oviducts of adult and senescent quails by the methylation method revealed that in the case of senescent animals an increase in shorter poly(A) tracts can be detected. These results support the assumption that the age-dependent reduction of avidin synthesis is also caused by changes in the control mechanisms on the post-transcriptional level.

Aging↗

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Measurement of the molecular weight distributions in human muscular deoxyribonucleic acid.

Highly purified DNA has been prepared from human muscle tissue and treated with three different single-strand specific desoxyribonucleases. The molecular weight of treated individual DNA molecules was determined by length measurements. Average DNA molecular weights and their standard deviations were correlated to the age of donors. No significant correlation could be established using seven different mathematical functions. Factor analytical considerations suggested grouping according to extreme smoking habits. Two populations with significant double linear correlations of age versus molecular weight and standard deviation then emerge. From the data it may be speculated that in heavy smokers with increasing age (or time of smoking) high molecular weight DNA integrity is lost to an increasing extent in post-mitotic tissue.

Age Factors↗

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Different deoxyribonucleases in human submandibular saliva.

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Deoxyribonucleases↗

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A sensitive method for identification of DNA dependent DNA polymerases in acrylamide gels after seperation by micro disc electrophoresis.

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Acrylamides↗

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Characterization of different deoxyribonucleases in human lymphocytes.

Four groups of deoxyribonuclease activities from human lymphocytes have been characterized by deoxyribonuclease assay in DNA-containing polyacrylamide gels following their separation by disc-electrophoresis. All activities hydrolyse DNA endonucleolytically. One neutral deoxyribonuclease found in the cytoplasmic fraction prefers native or UV-irradiated DNA over denatured DNA as substrate and is a 5'-monoester former. Two groups of acid deoxyribonuclease activities are detectable in the nuclear fraction. Both are 3'-monoester formers. One is as well active with denatured DNA as with native DNA, the other one shows the same activity with native and UV-irradiated DNA but lower activity with denatured DNA. An alkaline deoxyribonuclease activity, also localized in the nucleus, is a 5'-monoester former, and prefers denatured or UV-irradiated DNA as substrate.

Cell Nucleus↗

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Fractionated precipitation of acid macropolyanions by dialysis, a simple method for the estimation of DNA in complex biological samples.

After efficient extraction by para-aminosalicylate, chopping, grinding and eventual sonication, the macropolyanions are transformed into their cetyltrimethylammonium salts. These have differing solubilities, strongly depending on ionic strength. The cationic detergent-macropolyanionic salts are solubilized by high salt concentration. Salt is then dialysed out, rendering the polyanions highly insoluble in a sequential fashion. The insolubilized components are determined quantitatively by monitoring turbidity, which in case of DNA is strictly proportionate to its concentration. This relation is not affected by other components. This makes DNA determination possible even in crude aqueous extracts. The method has been applied to different objects, such as bacteria, plants, animals, soil and activated sludge. The method may prove to be especially useful in research of environmental poisons e.g. in rivers, lakes or clarifiers.

Animals↗

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An isocratic high-pressure liquid chromatographic purification method for radioactively labeled deoxyribonucleoside triphosphates.

A method is described for the rapid purification of radioactively labeled deoxyribonucleoside triphosphates from their spontaneously emerging hydrolysis products deoxyribonucleoside diphosphate, deoxyribonucleoside monophosphate, and deoxyribonucleoside. The separations which are finished within 3 min or less are carried out on a 0.1 X 5 cm column filled with LiChrosorb-NH2, using isocratic elution with 0.025 M potassium phosphate, pH 6.8, in a high-pressure liquid chromatograph at room temperature and a flow rate of 30 ml-h-1 (flow velocity 63.7 cm-min-1).

Chromatography, High Pressure Liquid↗

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The determination of the DNA base composition in 19 species of adriatic sponges with high-pressure liquid cation-exchange chromatography.

The (adenine+thymine)/(guanine+cytosine) base ratios of 19 species of adriatic sponges have been determined by high-pressure liquid cation-exchange chromatography. The base ratios vary from 1.49 (Mycale massa) to 0.63 (Hippospongia communis) according to an (A+T) content of 59.7 and 38.6 mol%, respectively. The DNAs of sponges of the order Keratosa showed marked differences in their (A+T) contents (39.5 to 58.8 mol%) whereas those of Tetractinellida and Halichondrina were nearly identical (39.3 to 40.8 and 49.5 to 49.8 mol%, respectively). The 5-methylcytosine (5MC) content was determined in 8 sponge DNAs by a semiquantitative method. The values differed from 0.8 to 2.2 mol% of 5MC.

Animals↗

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Deoxyribonucleases in herpes simplex virus type 1 and 2 infected primary rabbit kidney cells.

In primary rabbit kidney cells infected with herpes simplex virus four different neutral deoxyribonuclease activities can be detected by means of the deoxyribonuclease assay in DNA-containing polyacrylamide gels following their separation by disc-electrophoresis. The method is suitable to follow independently the change in each activity of the different enzymes using only about 5 X 10(5) cells for each assay during the time-course of infection. Under these conditions one enzyme activity is constant, two disappear while the activity of a fourth one present only in infected cells, increases.

Animals↗

PubMedDate not supplied

[Different deoxyribonuclease-activities in bull seminal plasma (author's transl)].

By means of the in situ assay of deoxyribonucleases in DNA-containing polyacrylamide gels after separation by micro-disc-electrophoresis different deoxyribonucleases are detectable in bull seminal plasma. There are two groups of acid deoxyribonuclease-activities with a pH optimum at pH 5.0, one with a pH optimum at pH 7.4 and an additional one with a pH optimum at pH 8.5.

Animals↗