Low Molecular Weight RNA Components Research Articles

Effect of 5,6-dichloro-1-β-D-ribofuranosylbenzimidazole on the synthesis of low molecular weight RNA components

The synthesis of low molecular weight RNA components D, C and A is more inhibited by 5,6-dichloro-1-β-D-ribofuranosylbenzimidazole than the synthesis of ribosomal RNA, which in turn is more inhibited than that of tRNA and 5S RNA. The differential inhibitory effect of 5,6-dichloro-1-β-D-ribofuranosylbenzimidazole is most pronounced at low concentrations. The results support the conclusion that D, C and A are products of polymerase II, and suggest that the D, C and A transcripts are derived from large size heterogeneous nuclear RNA. Components K and L are inhibited like tRNA and 5S RNA and are thus probably synthesized by polymerase III.

Small RNA species in maize tissue

The low molecular weight RNA components of maize have been analyzed after labeling callus and leaf tissue with [(3)H]uridine in vitro. Electrophoresis of the isolated RNA on acrylamide slab gels reveals, apart from 5S and transfer RNA, three major and about five minor RNA species with chain lengths between 140 and 280 nucleotides. These RNA molecules are labeled as rapidly as 5S, transfer RNA, and do not represent degradation products of large ribosomal RNA molecules. Furthermore, like 5S and transfer RNA, these small RNA species are stable and show no detectable turnover within forty-eight hours. Fractionation of the tissue into crude subcellular fractions indicates a preferential association of some of the small stable RNA species with the nucleus, while others appear to be located in the cytoplasm. The low molecular weight RNA spectrum from the leaf is similar to that observed in callus, with the major small RNA species equally present in both tissues.

Synthesis of low molecular weight RNA components in cells with a temperature-sensitive polymerase II

AF 8 cells are a mutant cell line of baby hamster kidney cells with a temperature-sensitive polymerase II activity. When these cells grow at the non-permissive temperature (40° C) the synthesis of low molecular weight RNA components D, C and A is preferentially inhibited, whereas the synthesis of rRNA, tRNA, 5 S RNA and component L is affected only a little or not at all. These results indicate that polymerase II catalyzes the synthesis of components D, C and A.

Synthesis of low molecular weight RNA components A, C and D by polymerase II in α-amanitin-resistant hamster cells

In an attempt to establish which RNA polymerase catalyzes the synthesis of the low molecular weight RNA components A, C and D, Ama 1 cells (mutant Chinese hamster cells) were used in experiments with addition of alpha-amanitin. Ama 1 cells contain an altered RNA polymerase II which is 800 times more resistant towards inhibition by alpha-amanitin than the wild type enzyme. Alpha-amanitin (up to 200 microgram/ml) added to these cells does not affect the synthesis of the low molecular weight RNAs A, C and D. These data together with our previous data showing that alpha-amanitin (0.5 - 5.0 microgram/ml) preferentially inhibits the synthesis of A, C and D in normal cells indicate that RNA polymerase II catalyzes the synthesis of the low molecular weight RNA components A, C and D.

The differential inhibitory effect of α-amanitin on the synthesis of low molecular weight RNA components in BHK cells

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Occurrence and properties of low molecular weight RNA components from cells at different taxonomic levels

1. 1. The occurrence and gel electrophoretic properties of low molecular weight RNA components (LMW RNA) have been studied in species at different taxonomic levels. The LMW RNA components apart from tRNA, 5S RNA and 5.5S RNA are called LMW∗RNA. 2. 2. The major components of LMW∗RNA in mammalian cells are L, A, C and D, accounting for 0.1–0.7% of cellular RNA. The gel electrophoretic migration of components L, C, and D is similar in different mammals but the migration of component A shows differences. 3. 3. Amphibia, reptiles and birds contain L, A, C and D in about the same amounts as mammals but slight differences in migration are seen for L, C and D. Component A is absent from the nucleated red blood cells of the chicken and the frog. 4. 4. Sea urchins contain three LMW∗RNA components with migrations different from L, A, C and D. These components account for about 0.1% of the cellular RNA. 5. 5. Insects contain only one LMW∗RNA component, migrating as component L. 6. 6. Tetrahymena, Physarum and Mycoplasmas have one component which may be a counterpart to component L in higher cells. Yeast shows no LMW∗RNA components. 7. 7. In the multicellular species the occurrence and gel electrophoretic migration of LMW∗RNA components are not related to tumorgenicity, developmental stage or origin of tissue.

Influence of mycoplasma infection on the incorporation of different precursors into RNA components of tissue culture cells

Seven different tissue culture cells have been cultured with and without mycoplasma ( M. hyorhinis) in the presence of various precursors of RNA. Total cellular RNA was isolated and analysed by electrophoresis on polyacrylamide gels. The results obtained with mycoplasma-infected cells can be summarized as follows: 1. 1. When cells are labelled with [8- 3H]guanosine or [5- 3H]uridine there is some incorporation into host cell 28S and 18S rRNA, but it is less than into mycoplasma 23S and 16S rRNA. [8- 3H]guanosine or [5- 3H]uridine are also incorporated into host cell and mycoplasma tRNA and mycoplasma 4.7S RNA, but the incorporation into host cell 5S rRNA and low molecular weight RNA components (LMW RNA) is reduced. 2. 2. [5- 3H]uracil is not incorporated into host cell RNA but into mycoplasma tRNA, 4.7S RNA, a mycoplasma low molecular weight RNA component M 1 and 23S and 16S rRNA. 3. 3. [ 3H]methyl groups are incorporated into mycoplasma tRNA, 23S and 16S rRNA, but not into host cell 28S, 18S, 5S rRNA nor into mycoplasma 4.7S RNA. 4. 4. With [ 32P]orthophosphate or [ 3H]adenosine as precursors, the labelling is primarily in the host RNA. Mycoplasma infection influences the labelling of RNA primarily by an effect on the utilization of the exogenously added radioactive RNA precursors, since the generation time of mycoplasma infected cells is about the same as that of uninfected cells. Mycoplasma infection may completely prevent the identification of LMW RNA components.

Einbau von 3H-Uracil in die RNA steriler Anzuchten von Utricularia stellaris mit und ohne Rohrzucker

Sucrose has been shown to increase the growth of Utricularia stellaris in axenic culture (Harder and Zemlin, 1967). Comparable cultures of Utricularia were incubated for 3 and 7 hours with tritiated uracil, and then nucleic acids were prepared. With sucrose, the incorporation is about 5 times higher than that with the inorganic salt cultures. After chromatographic separation of the nucleic acids on MAK-columns, different peaks of short time labelled RNA with high specific activity were found. These peaks indicate the presence of two low molecular weight RNA components in the inorganic culture. These peaks also have been found in cultures with sucrose, but in these cultures there is another predominant peak of high molecular weight RNA which is eluted from the column after the greater ribosomal RNA component.

Low molecular weight RNA components from KB cells.

Simple oligonucleotide maps can be obtained by two-dimensional paper electrophoresis from enzyme digests of 5S RNA from KB carcinoma ribosomes. These maps can be distinguished from those obtained from 5S RNA derived from E. coli and from RNA components of low molecular weight derived from KB cells with and without adenovirus infection.

Disc electrophoresis of ribonucleic acid in polyacrylamide gels

Zone electophoresis in polyacrylamide gels has been found to afford a sensitive method for the analysis of low molecular weight RNA components, notably soluble RNA. The electrophoretic mobilities of RNA components depend on their molecular weights and have been correlated with their sedimentation coefficients. The separation is substantially independent of pH, provided this does not approach a pK of the constituent nucleotides; it is also unaffected by the presence of urea. The effect of gel concentration and other experimental factors in the separation have been investigated and optimum conditions for any given molecular weight range within the scope of the method have been determined. Techniques are described for the quantitative estimation of the proportions of RNA components in a mixture. Using the disc electrophoresis procedure, analyses can be performed on less than 200 μg of RNA. Components may be almost quantitatively extracted from the gel after electrophoresis. Results for various sRNA preparations are given. All samples so far examined have many components of both lower and higher molecular weight than the main portion; these generally amount in all to some 25–30 percent of the total RNA in the preparation.