Analysis of Small RNA Species: Phylogenetic Trends
S. K. Dutta in DNA Systematics, 2019
A small stable 10S RNA has been identified and characterized from E. coli.121 Ribonuclease P, unique among all the RNA processing enzymes, contains an RNA moiety that is required for its function.122,123 Two RNAs are present in this enzyme. One of them, termed M2, is identical or similar to 10S RNA from E. coli. Also, ribonuclease P from yeast and Bacillus subtilis contains both protein and RNA components.124 Using a cloned segment of DNA for complementation it has been shown that it codes for an RNA species of 340 bases. Sequence analysis of RNA indicates that this RNA is highly G + C rich.125 It originates from DNA. It would be interesting to determine if GC rich 5.5S RNA from Alcaligenes faecalis originates from 10S RNA bound to RNAse.
“Biologically Active” RNA and the Immune Response*
Edward P. Cohen, A. Arthur Gottlieb in Immune RNA, 2020
Cohen’s extracts, as well as Fishman’s, were impure and contained, in addition to RNA, both DNA and protein. However, the likely essential participation (if not elucidated) of RNA in the response was indicated by the extraordinary sensitivity of the active extracts to ribonuclease. Enzyme:substrate ratios as high as 1:700,000 inactivated the extracts (approximately 0.001 μg ribonuclease destroyed the biological activity of 700 μg of extract dissolved in saline in less than 10 min at 20°). Extracts dissolved in medium containing divalent cations were partially resistant to ribonuclease. In later studies Cohen found18 that divalent cations aid in stabilizing the secondary structure of the RNA (Figure 1); this is indicated by the heightened hyperchromicity of RNA dissolved in buffer containing calcium and magnesium relative to that observed for aliquots of the RNA solution dissolved in buffer without calcium and magnesium. This observation deserves special emphasis. Later sections of this book will present the capacity of RNA extracts injected into immunocompetent recipients, including humans, to affect the immune status of the recipient. Ribonuclease is present in blood and intercellular fluids and would be expected to quickly degrade such RNA. RNA dissolved in solutions containing salts in physiologic concentrations resisted degradation. Persistence of its biological activity was noted after incubation with ribonuclease if the reaction was performed in medium containing calcium and magnesium, but not if the reaction was performed in medium free of divalent cations.
Hormonal Regulation of Gene Transcription
Gerald M. Kolodny in Eukaryotic Gene Regulation, 2018
In evaluating this work to date, there seems to be some reluctance to define in vitro effects in precise biochemical terms, particularly in work using crude cytosol preparations. An evaluation of the various subreactions of RNA synthesis and enzyme activities capable of modifying the chromatin DNA template or the RNA product is needed. Receptor preparations do contain ribonuclease activity.182 In other cases, the nature of the enhanced activity requires further definition; for example, effects in the rat prostate system are α-amanitin insensitive,176 but whether this represents Pol I, Pol III, or ribohomopolymer formation is not clear; as pointed out by Buller et al.,183 contamination of receptor and enzyme preparations with nucleic acid synthetases is a real problem. Controls and subcontrols are not always satisfactory; in one report,174 a statement to the effect that cytosol (without hormone) did not modify Pol activity is difficult to believe in the light of another report82 indicating that cytoplasmic extracts per se inhibit RNA synthesis. Dose-response curves, data on reproducibility, and information on the state of the Η-R complex during the incubation are also required. In the latter case, Davies and Griffiths’75 demonstrate the conversion of 8S receptor to 3S or 4 to 4.5S forms prior to association of the latter with chromatin; however, receptor dimers appear to be more effective in inducing in vitro responses in both chick oviduct181 and rat prostate systems.175
Novel RNASET2 Pathogenic Variants in an East Asian Child with Delayed Psychomotor Development
Published in Fetal and Pediatric Pathology, 2018
Yan Sun, Xuyun Hu, Jiqing Song, Yanyan Hu, Caihong Liu, Guimei Li
Ribonuclease T2 is the only member of the Rh/T2/S family of acid ribonuclease. Its function is involved in lysosomal degradation of ribosomal RNA. Its presence in locations where there is no RNA suggests that this ribonuclease may have other functions (9). Subsequent studies revealed its antiangiogenic and antitumorigenic effects independent of its ribonuclease capacity (9–12). Based on this feature, human recombination RNASET2 has been proposed as a potential anticancer agent in recent studies (12,14–16). Moreover, genome-wide association study (GWAS) results also revealed relations between Single Nucleotide Polymorphisms (SNPs) on RNASET2 and Graves' disease, Crohn's disease, rheumatoid arthritis, vitiligo and other autoimmune diseases (17–25). RNASET2 mutation was also reported in patients with CLWM and AGS. The distinctive findings of CLWM are bilateral temporal lobe cysts combined with a specific pattern of multifocal white matter lesions, normo- or microcephaly, and severe psychomotor retardation in a nonprogressive clinical course, while AGS is another inherited leukodystrophy with cerebral calcification. Nevertheless, the underlining basis between RNASET2 mutation and congenital brain infection-like phenotypes is still not clear.
Design, synthesis and biological evaluation of novel diarylpyridine derivatives as tubulin polymerisation inhibitors
Published in Journal of Enzyme Inhibition and Medicinal Chemistry, 2022
Shanbo Yang, Chao Wang, Lingyu Shi, Jing Chang, Yujing Zhang, Jingsen Meng, Wenjing Liu, Jun Zeng, Renshuai Zhang, Yingchun Shao, Dongming Xing
HeLa cells were inoculated in 6-well plates at a density of 2 × 105 per well and grown for 24 h. Cells were treated with 1-fold IC50, 2-fold IC50 and 3-fold IC5010t for 24 h. Cells were collected by centrifugation, washed with PBS and fixed overnight in ice-cold 70% ethanol. The fixed cells were collected by centrifugation and 100 μL of ribonuclease (RNase) was added. After a water bath at 37 °C for 30 min, 400 μL PI staining was added and the samples were stained for 30 min in the dark at 4 °C. Finally, the samples were analysed by flow cytometry (Beckman Coulter, USA). The data were processed and evaluated using software.30,31
Review of Chlamydia trachomatis viability methods: assessing the clinical diagnostic impact of NAAT positive results
Published in Expert Review of Molecular Diagnostics, 2018
Kevin J. H. Janssen, Jeanne A. M. C. Dirks, Nicole H. T. M. Dukers-Muijrers, Christian J. P. A. Hoebe, Petra F. G. Wolffs
However, due to its chemically labile composition, working with mRNA as a template can be challenging. RNA is more susceptible for degradation by heat than DNA and should be kept on ice during the complete workflow. Moreover, RNA can be degraded easily by ribonuclease (RNAse) activity, thus the use of RNAse inhibitors is highly recommended [64]. In addition, caution must be taken when interpreting results, as RNA purification methods do not always completely eliminate genomic DNA which can result in false positive results. Furthermore, there is no consensus on how long mRNA persists after chlamydia death, and how much this differs between different transcripts [53].
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