Evidence for complete symmetrical transcription in vivo of mitochondrial DNA in HeLa cells.
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Biomedical subjects
Publications and source records attributed to C Tu.
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The selective incorporation of 5-bromodeoxyuridine (BrdU) into mitochondrial DNA (mit-DNA) in the LM(TK-) ClID cell line, a thymidine kinase-deficient derivative of L fibroblasts with persistent mitochondrial enzyme activity, has been utilized to specifically damage mit-DNA by 'visible' light irradiation. ('Visible light' indicates the source of light used, although the components most active photochemically on BrdU-substituted DNA are in the near-visible range, 300-340 nm.) (Szybalski & Opara-Kubinski, 1965). LM(TK-) Cl ID cells, which had been grown in the presence of 30 mug/ml BrdU, were irradiated with 'visible' light. Analysis of the pre-existing mit-DNA in these cells, which had been long-term labelled with [5-3H]deoxycytidine, showed a progressive decrease, with increasing duration of irradiation, in the proportion of the closed-circular form and an increase in that of the open-circular form of mit-DNA, with the subsequent appearance of fragments of this DNA. Furthermore, there was a decrease during irradiation in the total amount of mit-DNA, which became about 35% of the non-irradiated control after 65 h irradiation. On the other hand, irradiation with 'visible' light failed to cause any quantitative or qualitative change, with respect to the non-irradiated control, in mit-DNA from cells grown in the absence of BrdU and long-term labelled with [Me-3h]thymidine. An analysis of the incorporation of [5-3H]deoxycytidine into mit-DNA of BrdU-grown cells, during a 3-h exposure of the cells to the precursor following irradiation, showed a fairly rapid decline of mit-DNA labelling; this became about 50% of the non-irradiated control after 12 h irradiation, decreasing to about 25% in the next 48 h. By contrast, no effect of irradiation was observed on the subsequent pulse-labelling of mit-DNA with [Me-3H]thymidine in cells grown in the absence of BrdU. Furthermore, no change in the size of the extracted nuclear DNA was found in irradiated BrdU-grown cells. The progressive and selective damage and destruction of mit-DNA during irradiation with 'visible' light of Cl ID cells correlate fairly well with the kinetics of loss of cell viability occurring under the same conditions, as described in the accompanying paper, strongly suggesting a link between the two phenomena.
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Protein has been shown to be the principal source of leucine for the Day 8.5 to 10.5 rat conceptus in culture. It could be argued that this finding applies only after adaptation to culture conditions and does not apply in vivo. This possibility was investigated using an isotope-dilution technique after i.v. injection of [3H]leucine into Day 10.5 pregnant rats. Specific radioactivity of free leucine in the conceptus was 8 to 10% of that in maternal plasma. Slow exchange of leucine with the maternal circulation and fetal tissue protein turnover were judged to be inadequate as explanations for the observed isotope dilution. Taken together, our results and those from in vitro studies are consistent with a major contribution of leucine coming from the degradation of protein in vivo, probably involving the visceral yolk sac. Our results suggest that mechanisms of amino acid supply to the conceptus identified using whole-embryo culture mirror those in vivo.
A new approach, based on a histogram and genetic algorithm (GA), has been developed to rapidly and reliably detect QRS complexes of the ECG curve. Here, the new approach is described and the principle of the histogram and GA is explained. The results of detection of QRS complexes using the new method are given. Additionally, as an example of its novel application, the P-wave and f-wave (when atrial fibrillation is absent and present, respectively) are extracted easily and efficiently from the ECG curve using the method.