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PubMedDate not supplied

How sleep and mental disorders are related to complaints of daytime sleepiness.

BACKGROUND: Daytime sleepiness is widespread and has negative impacts on the public sector. OBJECTIVE: To ascertain the incidence and prevalence of daytime sleepiness and associated risk factors in the general population. METHOD: In 1994, a representative sample of the non-institutionalized British population aged 15 years or older was interviewed via telephone using an expert computer-assisted program designed to facilitate surveys of this type (Sleep-Eval, M. M. Ohayon, Montreal, Quebec). Subjects were classified into 3 groups based on the severity of their daytime sleepiness. We completed 4972 interviews (acceptance rate, 79.6%). RESULTS: Severe daytime sleepiness was reported in 5.5% (95% confidence interval, 4.9%-6.1%) of the sample, and moderate daytime sleepiness in another 15.2% (95% confidence interval, 14.2%-16.2%). Associated factors with severe daytime sleepiness included female sex, middle age, napping, insomnia symptoms, high daily caffeine consumption, breathing pauses or leg pain in sleep, depressive disorder (based on the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition, criteria), falling asleep while reading or watching television, and motor vehicle crashes or accidents involving use of machinery. Moderate daytime sleepiness was associated with female sex, napping, insomnia symptoms, arthritis or heart disease, and gross motor movements during sleep. CONCLUSIONS: It is likely that daytime sleepiness deleteriously affects work activities, social and/or marital life, and exhibits a negative socioeconomic impact. In addition, the risk of a motor vehicle crash appears to be higher in this specific population: twice as many subjects operating a motor vehicle or using machine tools reported having a crash or accident, respectively, in the previous year in the groups with severe daytime sleepiness or moderate daytime sleepiness than did the general population with no daytime sleepiness. The high prevalence rates of daytime sleepiness and multiplicity of related factors mandate further scrutiny by public health officials.

Adolescent↗
PubMed2004

Tissue- and stage-specific RNA editing of rps 14 transcripts in moss (Physcomitrella patens) chloroplasts.

The C to U editing event that converts an ACG codon to an AUG translation initiation codon in the chloroplast rps 14 transcript is unique to the moss Physcomitrella patens and has not been found in other species. The efficiency of RNA editing was 80% in the young protonemata and decreased to approximately 20% in old protonemata and fully developed leafy shoots. This indicates that RNA editing of this site is regulated in a tissue- and stage-specific manner. In this study, a novel C to U RNA editing site has been identified at the -1 position relative to the AUG. Because the editing site is localized in the mRNA 5' untranslated region, it may affect the efficiency of rps 14 mRNA translation.

Base Sequence↗
PubMed2000

APOBEC-1 dependent cytidine to uridine editing of apolipoprotein B RNA in yeast.

Cytidine to uridine editing of apolipoprotein B (apoB) mRNA requires the cytidine deaminase APOBEC-1 as well as a tripartite sequence motif flanking a target cytidine in apoB mRNA and an undefined number of auxiliary proteins that mediate RNA recognition and determine site-specific editing. Yeast engineered to express APOBEC-1 and apoB mRNA supported editing under conditions of late log phase growth and stationary phase. The cis -acting sequence requirements and the intracellular distribution of APOBEC-1 in yeast were similar to those described in mammalian cells. These findings suggest that auxiliary protein functions necessary for the assembly of editing complexes, or 'editosomes', are expressed in yeast and that the distribution of editing activity is to the cell nucleus.

APOBEC-1 Deaminase↗
PubMed1992

The solution structure of a cyclic endothelin antagonist, BQ-123, based on 1H-1H and 13C-1H three bond coupling constants.

A cyclic pentapeptide endothelin antagonist, cyclo(dTrp-dAsp-Pro-dVal-Leu), recently reported (K. Ishikawa et al., 13th Am. Pept. Symp., Cambridge MA, 1991) has been studied by NMR spectroscopy and molecular modeling. A stable structure has been determined without the use of nuclear Overhauser effects and is based primarily on homonuclear and heteronuclear three bond coupling constants. The 13C-edited TOCSY experiment is demonstrated at natural abundance and approximately 30 mM peptide concentrations. Three bond 13C-1H coupling constants obtained by this method are shown to reduce the ambiguity in phi angle determination which exists when only interproton coupling constants are used. Three out of four phi angles were determined uniquely by this method and the fourth was reduced to two possible values. The proline phi angle was determined to be -78 degrees based on the 3JH alpha, H beta and 3JH alpha, H beta coupling constants. Comparison of amide proton temperature dependence, chemical shifts and vicinal proton coupling constants in a 20% acetonitrile/80% water solvent mixture and in (CD3)2SO indicates that the structure is similar in both solvents.

Amino Acid Sequence↗
PubMed1996

A duplicated sequence in sugarbeet mitochondrial transcripts is differentially edited: analysis of orfB and its derivative orf324 mRNAs.

RNA editing of the duplicated sequences was investigated in the transcripts of orfB and orf324 genes from sugarbeet mitochondria. The orf324 shares the first 59 bp of the reading frame and 321 bp upstream sequence with orfB. Two cytidine-to-uridine editing sites were found in orfB, but the corresponding cytidine residues remained unchanged in the transcripts of orf324. In the vicinity of the non-edited cytidine residues within the shared sequence element of orf324 were identified three point mutations that may abolish orf324 editing. Our results also suggest that selection of editing sites depends on primary sequence.

Amino Acid Sequence↗
PubMed1991

Cycles of progressive realignment of gRNA with mRNA in RNA editing.

We characterized numerous partially edited NADH dehydrogenase 7 and ATPase 6 cDNAs. Most of these have a stretch of incompletely edited sequence at the junction of mature and unedited sequences. The characteristics of the junctions suggest editing of sites multiple times and that editing within each junction does not proceed precisely 3' to 5'. Analyses of gRNAs and corresponding junction sequences predict a series of progressively more stable, but incompletely base-paired, interactions in the junction region. The predicted interactions suggest that the gRNA is progressively realigned with the mRNA being edited. We suggest that gRNA interactions with the mRNA result in regions of lower thermodynamic stability that are selected for editing, thus driving toward the most stable structure, the complete gRNA/mRNA duplex.

Adenosine Triphosphatases↗
PubMed1997

Insertional editing of nascent mitochondrial RNAs in Physarum.

Maturation of Physarum mitochondrial RNA involves the highly specific insertion of nonencoded nucleotides at multiple locations. To investigate the mechanism(s) by which this occurs, we previously developed an isolated mitochondrial system in which run-on transcripts are accurately and efficiently edited by nucleotide insertion. Here we show that under limiting concentrations of exogenous nucleotides the mitochondrial RNA polymerases stall, generating a population of nascent RNAs that can be extended upon addition of limiting nucleotide. Several of these RNA species have been characterized and were found to be fully edited, indicating that nascent RNA is a substrate for nucleotide insertion in isolated Physarum mitochondria. Remarkably, these RNAs are edited at positions located within 14-22 nucleotides of the polymerase active site, suggesting that insertional editing may be physically or functionally associated with transcription. The absence of unedited RNA in these experiments indicates that large tracts of RNA downstream of editing sites are not required for nucleotide addition, and argues that insertional editing in Physarum occurs with a 5' to 3' polarity. These data also provide strong evidence that insertional editing in Physarum is mechanistically distinct from editing in kinetoplastid systems.

Animals↗
PubMed1995

The secondary structure of guide RNA molecules from Trypanosoma brucei.

RNA editing in kinetoplastid organisms is a mitochondrial RNA processing phenomenon that is characterized by the insertion and deletion of uridine nucleotides into incomplete mRNAs. Key molecules in the process are guide RNAs which direct the editing reaction by virtue of their primary sequences in an RNA-RNA interaction with the pre-edited mRNAs. To understand the molecular details of this reaction, especially potential RNA folding and unfolding processes as well as assembly phenomena with mitochondrial proteins, we analyzed the secondary structure of four different guide RNAs from Trypanosoma brucei at physiological conditions. By using structure-sensitive chemical and enzymatic probes in combination with spectroscopic techniques we found that the four molecules despite their different primary sequences, fold into similar structures consisting of two imperfect hairpin loops of low thermodynamic stability. The molecules melt in two-state monomolecular transitions with Tms between 33 and 39 degrees C and transition enthalpies of -32 to -38 kcal/mol. Both terminal ends of the RNAs are single-stranded with the 3' ends possibly adopting a single-stranded, helical conformation. Thus, it appears that the gRNA structures are fine tuned to minimize stability for an optimal annealing reaction to the pre-mRNAs while at the same time maximizing higher order structural features to permit the assembly with other mitochondrial components into the editing machinery.

Animals↗
PubMed1997

Guide RNA requirement for editing-site-specific endonucleolytic cleavage of preedited mRNA by mitochondrial ribonucleoprotein particles in Trypanosoma brucei.

RNA editing in trypanosome mitochondria entails the posttranscriptional internal addition and occasional deletion of uridines from precursor mRNAs. Ample evidence exists to show that the information specifying the site and number of uridines added or deleted comes from small, mitochondrially encoded guide RNAs (gRNAs). More recent work indicates that the process involves an enzymatic cascade, initiating with an endonucleolytic cleavage of the pre-mRNA at an editing site. The cleaved editing site can undergo uridine (U) addition to or deletion from the 3' end of the 5' fragment via a mitochondrial terminal uridylyl transferase (TUTase) or terminal uridylyl exonuclease, respectively. Mitochondrial RNA ligase subsequently rejoins the mRNA. Activities to carry out these processes have been found in trypanosome mitochondria, including an editing-site-specific endonuclease activity which cleaves preedited but not edited mRNAs. We have found that this enzymatic activity cosediments with the same 19S ribonucleoprotein particle previously shown to contain TUTase, RNA ligase, and gRNAs and remains stable after salt treatment. Depletion of endogenous cytochrome b gRNAs by the addition of complementary oligonucleotides in vitro completely inhibits editing-site cleavage of synthetic preedited cytochrome b mRNA. The addition of synthetic cognate gRNA for cytochrome b but not unrelated small RNA restores editing-site cleavage. These studies show that in addition to specifying the site and number of uridines added or deleted, gRNAs provide the necessary information for cleavage by the editing-site-specific endonuclease.

Adenosine Triphosphate↗
PubMed2000

Development. Survival is impossible without an editor.

RNA editing is a fascinating phenomenon that is found in both animal and plant cells. By converting an adenosine base to an inosine (which behaves like guanosine) in RNA that has already been transcribed, certain RNA sequences (and hence the amino acids they encode) are altered. In a Perspective, Keegan, Gallo and O'Connell explore new results showing that activity of the editing enzyme ADAR1 is crucial for normal development of red blood cells in mouse embryos.

Adenosine↗
PubMed2000

DNAssist: the integrated editing and analysis of molecular biology sequences in windows.

MOTIVATION: The programs currently available for the analysis of nucleic acid and protein sequences suffer from a variety of problems: Web-based programs often require inconvenient reformatting of sequences when proceeding from one analysis to the next, and commercial-console-based programs are cost prohibitive. Here, we report the development of DNASSIST:, an inexpensive, multiple-document, interface program for the fully integrated editing and analysis of nucleic acid and protein sequences in the familiar environment of Microsoft Windows.

Animals↗
PubMed1995

Regional differences in the extent of RNA editing of the glutamate receptor subunits GluR2 and GluR6 in rat brain.

The extent of RNA editing of the glutamate receptor subunits GluR2 and GluR6 was studied by using a newly developed method based on the restriction analysis of the subunit-specific polymerase chain reaction (PCR) product with the enzyme Bbv 1. Total RNA was isolated from following brain regions: cortex, striatum, hippocampus, thalamus, hypothalamus, cerebellum, pons/medulla oblongata and white matter. RNA was transcribed into cDNA, which was used as template for PCR. PCR was run with GluR2- and GluR6-specific primers to amplify a product across the edited region. The PCR products were analysed with the restriction enzyme Bbv 1 and gel electrophoresis of the restriction digest. Bbv 1 recognizes the sequence GCAGC which is identical with the sequence of the PCR product originating from unedited GluR2 or GluR6 mRNA. Thus, this enzyme splits the non-edited PCR product into two fragments while leaving the edited PCR product intact. After electrophoresis of the restriction digest and photographing gels, optical density of bands was quantified with image analysis. For quantification calibration curves were made with PCR products from constructs originating from edited and non-edited GluR6 mRNA. GluR2 mRNA was completely edited in all brain structures studied. Editing of GluR6 mRNA, in contrast, was high in gray matter structures (above 90%) but considerably lower in the pons/medulla oblongata (66%) and white matter (55%). It is, therefore, suggested that editing of GluR2 and GluR6 mRNA is performed by different enzymatic activities. Studying RNA editing of glutamate receptor subunits will extend knowledge about the role of calcium fluxes through non-NMDA glutamate receptor ion channels.

Animals↗
PubMed2025

A base editor facilitates simultaneous purine and pyrimidine substitutions for ex vivo and in vivo mutagenesis screens.

Genetic mutations are closely linked to human diseases, yet the relationship between many mutations and their corresponding phenotypes remains poorly understood. Furthermore, tools to study the connection between nucleotide variations and phenotypes are limited. To address this issue, we developed ACGBEmax by fusing the dual-functional deaminase, engineered N-methylpurine DNA glycosylase, and evolved SOS response associated peptidase domain with nCas9(D10A). ACGBEmax enables the precise conversion of A, C, and G to other bases in mammalian cells, thereby generating an extensive range of base mutations types. We used ACGBEmax to generate HPRT variants, identifying mutations conferring resistance to 6-thioguanine. Additionally, we performed in situ mutagenesis of Ctnnb1 in mouse liver, identifying both known and potential oncogenic mutations. Our results prove that ACGBEmax is a powerful tool for generating a wide spectrum of mutation types at specific gene loci, highlighting its significant potential for applications in functional screening and the directed evolution of protein variants.

Animals↗
PubMed2004

Gene-targeted mice lacking the Trex1 (DNase III) 3'-->5' DNA exonuclease develop inflammatory myocarditis.

TREX1, originally designated DNase III, was isolated as a major nuclear DNA-specific 3'-->5' exonuclease that is widely distributed in both proliferating and nonproliferating mammalian tissues. The cognate cDNA shows homology to the editing subunit of the Escherichia coli replicative DNA polymerase III holoenzyme and encodes an exonuclease which was able to serve a DNA-editing function in vitro, promoting rejoining of a 3' mismatched residue in a reconstituted DNA base excision repair system. Here we report the generation of gene-targeted Trex1(-/-) mice. The null mice are viable and do not show the increase in spontaneous mutation frequency or cancer incidence that would be predicted if Trex1 served an obligatory role of editing mismatched 3' termini generated during DNA repair or DNA replication in vivo. Unexpectedly, Trex1(-/-) mice exhibit a dramatically reduced survival and develop inflammatory myocarditis leading to progressive, often dilated, cardiomyopathy and circulatory failure.

Animals↗
PubMed1994

RNA editing of a human glutamate receptor subunit.

AMPA receptors are comprised of individual subunits, and the divalent cation permeability of assembled AMPA receptors is determined by a single amino acid residue in the second transmembrane region of the GluR-B subunit. At this site, GluR-B subunits contain an arginine while other AMPA receptor subunits contain glutamine. Interestingly, the murine gene for GluR-B actually specifies a glutamine at the divalent cation permeability site. The appearance of arginine and not glutamine in the mature GluR-B protein is thought to be a result of RNA editing of the GluR-B messenger RNA. In that AMPA receptors are thought to mediate the bulk of fast excitatory signalling within the mammalian central nervous system, this process of RNA editing may play a pivotal role in normal neural function by mediating divalent cation permeability of AMPA receptors. Disruptions of RNA editing could lead to phenotypically altered AMPA receptors, with implications for pathogenic brain processes. We report that the human GluR-B gene sequence is also edited such that there is a difference between the human GluR-B gene and the complementary DNA (cDNA), as demonstrated both with allele-specific polymerase chain reaction (PCR) and restriction enzyme digestion of PCR products. Thus, as in the rodent brain, RNA editing of an AMPA receptor subunit appears to be an important process in the human brain. Disruptions of RNA editing may have neuropathological consequences.

Base Sequence↗
PubMed2000

Double-stranded RNA adenosine deaminases ADAR1 and ADAR2 have overlapping specificities.

Adenosine deaminases that act on RNA (ADARs) deaminate adenosines to produce inosines within RNAs that are largely double-stranded (ds). Like most dsRNA binding proteins, the enzymes will bind to any dsRNA without apparent sequence specificity. However, once bound, ADARs deaminate certain adenosines more efficiently than others. Most of what is known about the intrinsic deamination specificity of ADARs derives from analyses of Xenopus ADAR1. In addition to ADAR1, mammalian cells have a second ADAR, named ADAR2; the deamination specificity of this enzyme has not been rigorously studied. Here we directly compare the specificity of human ADAR1 and ADAR2. We find that, like ADAR1, ADAR2 has a 5' neighbor preference (A approximately U > C = G), but, unlike ADAR1, also has a 3' neighbor preference (U = G > C = A). Simultaneous analysis of both neighbor preferences reveals that ADAR2 prefers certain trinucleotide sequences (UAU, AAG, UAG, AAU). In addition to characterizing ADAR2 preferences, we analyzed the fraction of adenosines deaminated in a given RNA at complete reaction, or the enzyme's selectivity. We find that ADAR1 and ADAR2 deaminate a given RNA with the same selectivity, and this appears to be dictated by features of the RNA substrate. Finally, we observed that Xenopus and human ADAR1 deaminate the same adenosines on all RNAs tested, emphasizing the similarity of ADAR1 in these two species. Our data add substantially to the understanding of ADAR2 specificity, and aid in efforts to predict which ADAR deaminates a given editing site adenosine in vivo.

Adenosine Deaminase↗
PubMed2006

Hahnemann's experiments with 50 millesimal potencies: a further review of his casebooks.

BACKGROUND: A review of Hahnemann's clinical records at the Institute for History of Medicine of the Robert Bosch Foundation in Stuttgart shows that until the end of his life, Hahnemann continued to refine his clinical method, based on clinical cases. His "most perfected method" motivated him to write the sixth edition of the The Organon of the Healing Art, proposing solutions controlling the side effects he observed with repeated doses of homeopathic medicines. Unfortunately, this was published many years posthumously. The sixth edition of The Organon introduced the fifty-millesimal scale. OBJECTIVES: To identify the clinical cases treated with fifty-millesimal potencies and analyze Hahnemann's use of them. RESULTS: 1836 prescriptions of fifty-millesimal potencies were found, between 1837 and 1843 in three phases: initially sporadic; later compared regularly to centesimal dynamizations; and finally systematically. Thirty five medicines were identified in fifty-millesimal prescriptions, seven in potencies higher than 10 and only 3 (Sulphur, Mercurius solubilis and Rhus toxicodendron) used in the 30th degree. This accords with Haehl's information about the remedies in Hahnemann's case of fifty-millesimal potencies. CONCLUSIONS: Hahnemann probably decided to write the sixth edition, in 1840, to incorporate his latest experience with the repetition of potentized doses and periodically modified potencies. He must have revised it after February 1842 to include his latest findings with fifty-millesimal potencies in ascending degrees. Hahnemann's conception about the superiority of the fifty-millesimal in comparison with the centesimal dynamization was based on a significant number of experiments with the two scales.

Germany↗
PubMed1995

The application of numerical estimates of base calling accuracy to DNA sequencing projects.

During DNA sequencing projects one of the most labour intensive and highly skilled tasks is to view the original trace descriptions of gels and to adjudicate between conflicting readings. Given the current methods of calculating a consensus, the majority of the time employed in viewing traces and editing readings is actually devoted to making the poorer data fit the good data. We propose new consensus calculation algorithms that employ numerical estimates of base calling accuracy and which when used in conjunction with an automatic detector of contradictory data should greatly reduce the time spent checking and editing readings and hence improve DNA sequencing productivity.

Algorithms↗