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The metabolism of high-molecular-weight ribonucleic acid including polyadenylated species, in the developing rat brain.

ABSTRACT: High-molecular-weight RNA was isolated from rat brain at various times after the intracranial administration of [32P]Pi. The synthesis of 28S and 18S rRNA could be detected within 1h of the injection of the radioactive precursor and appeared to be more pronounced, relative to other high-molecular-weight RNA, in the brains of older rats compared with those of newborn rats. Polyadenylated RNA, representing most mRNA and their precursors, was isolated by chromatography on oligo(dT)-cellulose. The contribution of this polyadenylated RNA to total RNA synthesis was investigated in the cerebral cortex and the phylogenetically older brain stem at different stages in the development of the rats by using a 5h period of labelling as an arbitrary index of transcription. In the brain stem the proportion of labelled polyadenylated RNA comprised 27-30% of the total RNA. The corresponding values for the cortex decreased from 34% in newborn rats to 23% in 40-150-day-old rats. These data indicated that proportionately more polyadenylated RNA is synthesized in the cortex of the newborn than in the adult rat and that there is a progressive decrease in the synthesis of polyadenylated RNA relative to rRNA during development.

SUBMITTER: Berthold W

PROVIDER: S-EPMC1172732 | biostudies-other | 1976 Feb

REPOSITORIES: biostudies-other

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Nucleo-cytoplasmic relationships of high-molecular-weight ribonucleic acid, including polyadenylated species, in the developing rat brain.

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The metabolism of high-molecular-weight ribonucleic acid in hypothalamic and cortical regions of the developing female rat brain.

Project description:The regional metabolism of high-molecular-weight RNA in the developing female rat brain was investigated after the intracranial injection of [32P]P1. The synthesis of polyadenylated RNA relative to high-molecular-weight RNA was determined after oligo(dT)-cellulose chromatography of total cellular high-molecular-weight RNA labelled after 4h. In both hypothalamus and cortex this synthesis was significantly higher during the first 10 days post partum than at subsequent ages. In both regions apparently more mRNA is synthesized in the young. The ratio of the specific radioactivity of cytoplasmic high-molecular-weight RNA relative to that of the nucleus, measured after a 48 h period of labelling, was considered to be an index of the nucleocytoplasmic transport of newly synthesized RNA [Berthold & Lim (1976) Biochem. J. 154, 529--539]. In the cortex, nucleo-cytoplasmic RNA transport in rats aged up to 20 days was significantly higher than in older rats, with the maximal value being attained between 16 and 19 days post partum. In contrast, in the hypothalamus, nucleo-cytoplasmic transport of RNA was low during the neonatal period and comparable with that of the mature animal. However, there were two periods of increased transport at later stages of development, the first between 15 and 19 days post partum and the second between 25 and 29 days post partum. These prepubertal changes in the nucleo-cytoplasmic transport of RNA in the female hypothalamus during weeks 3 and 4 post partum are coincident with other reported changes occurring during sexual differentiation. Differences in the timing of the maturational changes of the two brain regions thus appear to be reflected in developmental changes in RNA transport.

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TERMINAL-SEQUENCE STUDIES OF HIGH-MOLECULAR-WEIGHT RIBONUCLEIC. THE REACTION OF PERIODATE-OXIDIZED RIBONUCLEOSIDES , 5'-RIBONUCLEOTIDES AND RIBONUCLEIC ACID WITH ISONIAZID.

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The effect of thioacetamide on the maturation of high-molecular-weight ribonucleic acid in tumour cells.

Project description:1. Although thioacetamide treatment of Krebs II ascites-tumour cells did not markedly affect the rate of RNA synthesis in vivo, it caused the formation of an unusual single-stranded RNA component sedimenting at approx. 26s. 2. The maturation process leading to the formation of methylated RNA was examined by following the kinetics of incorporation into RNA of radioactivity from [G-(3)H]uridine and l-[Me-(14)C]methionine. In treated and untreated tumour cells extensive methylation was observed, not only of the ribosomal RNA species, but also of their precursors, especially the precursor species sedimenting at 35s. 3. Evidence is also presented to suggest that methylation of low-molecular-weight RNA species occurs both in the nucleus and in the cytoplasm of these tumour cells. 4. Thioacetamide did not appear to have an effect on RNA methylation in vivo, and in thioacetamide-treated cells the 26s RNA accumulated within the nucleus, where it was methylated. 5. It is postulated that the 26s RNA is most likely to arise as a result of a fault in the scission process that gives rise to the ribosomal RNA components from their high-molecular-weight precursors.

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Terminal-sequence studies of high-molecular-weight ribonucleic acid. The 3'-termini of rabbit reticulocyte ribosomal RNA.

Project description:Sequences of the polynucleotide chains of RNA found in the large and small ribosomal subunits of rabbit reticulocytes have been determined from the 3'-end by use of periodate oxidation and condensation with [(3)H]isoniazid and by stepwise degradation. By these methods the hexanucleotide sequences have been found as -pGpUpUpUpGpU for the 28S RNA and -pGpUpCpGpCpU for the 6S RNA of the large ribosomal subunit and the octanucleotide sequence -pGpApUpCpApUpUpA for the 18S rRNA of the small ribosomal subunit. These sequences are present in at least 70% of all the RNA molecules and are discussed in relation to the specific cleavage of rRNA from its precursors and the role of multiple cistrons for rRNA in the DNA of higher organisms. The feasibility of using the method for longer sequence determinations is discussed.

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High-molecular-weight nuclear polyadenylate-containing ribonucleic acid isolated from the lactating guniea-pig mammary gland contains milk-protein messenger ribonucleic acid sequences.

Project description:1. Steady-state poly(A)-containing nuclear RNA was isolated from the lactating guinea-pig mammary gland and analysed by sucrose-gradient centrifugation under denaturing conditions. 2. Nucleic acid-hybridization studies demonstrated the presence of small amounts of high-molecular-weight RNA species containing milk-protein mRNA sequences sedimenting at 25S. These were not found in the post-nuclear supernatant, where milk-protein sequences sedimented only between 12S and 15S; these were also the predominant species in the nuclear fraction. 3. The results are consistent with the transcription of milk-protein genes as high-molecular-weight precursor RNA species, 3-4 times as large as the active mRNA species isolated from the post-nuclear fraction.

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Next generation sequencing identifies polyadenylated species of hTR

Project description:The paired-end next-generation sequencing of all hTR versions of less than 200 nucleotides in length was used to analyze the 3’ end distribution of hTR associated to Flag-tagged versions of PABPN1, hTERT and Dyskerin. In total, we obtained 2,716,342, 3,152,013, and 3,077,395 hTR-specific reads for the PABPN1, hTERT, and Dyskerin purifications, respectively. We found that the majority of polyadenylated telomerase RNA recovered in the PABPN1, hTERT, and Dyskerin purifications had poly(A) tails starting immediately after the annotated hTR 3’ end. However, a greater proportion of poly(A) tails were found on genome-encoded 3’-extentions in the PABPN1-bound fraction compared to the population of hTR that was copurified with hTERT and DKC1: 15.5% for PABPN1, 6.9% for hTERT, and 6.5% for Dyskerin. Notably, the population of polyadenylated telomerase RNA recovered with PABPN1 was significantly different in terms of poly(A) tail length compared to polyadenylated hTR retrieved in the hTERT- and Dyskerin-bound fractions (P-value=5.551e-16, Kolmogorov-Smirnov test), showing a tendency toward longer poly(A) tails in the PABPN1-bound fraction. The enrichment of telomerase RNA with long poly(A) tails in the PABPN1-bound fraction is consistent with a role of PABPN1 in a maturation pathway that depends on hTR 3’ end polyadenylation. Moreover, our results indicate that a fraction of hTERT and Dyskerin are associated with polyadenylated versions of hTR.

2015-10-21 | GSE74186 | GEO

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