Biomedical subjects
Publications and source records attributed to E Probst.
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Previous work has indicated that at least some of the genetic information required for the expression of sea urchin histone genes is recognized following injection of the gene repeat (h22) into Xenopus oocyte nuclei. The ability to elicit the expression of cloned genes and their sequence-manipulated counterparts is proving invaluable in analysing the molecular details of gene expression. Direct injection of such genes into Xenopus oocyte nuclei remains one of the simplest methods for obtaining such expression and a remarkable degree of transcriptional fidelity has been demonstrated using this system with RNA polymerase III genes, and to a lesser extent with rDNAs transcribed by RNA polymerase I. In the case of polymerase II genes there is ample evidence for coupled transcription-translation, but the degree of transcriptional fidelity involved may, as has recently been shown for the ovalbumin gene, be minimal. However, clearly if the oocyte is to be used to investigate transcriptional regulation of such genes, transcriptional fidelity defined as the production of correct RNA termii, rather than the production of 'functional mRNAs' (ref. 15), must pertain. Here we demonstrate such fidelity in the expression of all five Psammechinus miliaris histone genes comprising a repeat unit. However, we find large quantitative variations in the levels of synthesis of the individual correct termini and hence of the mRNAs. In addition to the mRNAs, species with no detectable counterparts in the sea urchin are generated off the coding strand, as are heterogeneous noncoding species.
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The electrophoretic separation of labeled "9S" histone mRNAs obtained from cleaving sea urchin polysomes was found at first to be highly unreproducible. It became evident that the secondary structure of the individual mRNAs had a greater effect on their relative electrophoretic mobilities than did their molecular weight differentials. We determined the parameters affecting electrophoretic mobility by the novel method of running the labeled polysomal RNA in slab gels across polyacrylamide and urea gradients. The initially complex and species-specific electrophoretic pattern could then, by a judicious choice of denaturing conditions, be simplified to yield five well defined classes of labeled mRNAs. Using optimal conditions for the separation of the RNA components, five messengers were isolated from Psammechinus embryos by preparative disc electrophoresis, four of which, after two electrophoretic separations, exhibited a unimodal distribution. Each of the mRNAs was translated in vitro, four of the five fractions promoting the synthesis of one major protein. The in vitro products were characterized by comparison of their electrophoretic mobilities with those of known sea urchin histones. It was thus possible to correlate individual mRNAs with specific histones. We propose that the five mRNAs designated a-e in order of decreasing electrophoretic mobility code for the histones H4, H2A, H2B, H3, and H1.
The relationship between ornithine decarboxylase (L-ornithine carboxylyase, EC 4.1.1.17) activity and DNA synthetic activity was studied in mouse epidermis. Interfollicular epidermis and hair follicles were investigated separately. It was found that, in hair follicles, the variations of DNA replicative activity, which are reflected in the cyclic growth of hair, are paralleled by corresponding changes in ornithine decarboxylase activity. In both interfollicular epidermis and hair follicles, stimulation of DNA synthetic activity by plucking of hair induced a rapid and marked increase in ornithine decarboxylase activity. The relationship of steady-state and induced ornithine decarboxylase activity to DNA synthetic activity was compared in hair follicles and interfollicular epidermis. A correlation between the activity of this enzyme and DNA replication was found thereby in each of these tissues.
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