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The ntrBC genes coding for the bacterial signal-transducing protein NtrB and the bacterial enhancer-binding protein NtrC of deep-sea piezophilic Shewanella violacea were cloned and their nucleotide sequences were analyzed. The conserved regions of NtrB and those of NtrC are well conserved in the case of the ntrBC products of S. violacea.
Correction of disturbed respiratory function and inhibition of progression of pulmonary inflammatory and obstructive processes is the aim of respiration rehabilitation in patients with chronic bronchitis (CB). It is known that rehabilitation in CB patients with spinal diseases meets with difficulties as dystrophic lesions of the spine entail its functional-anatomic disorders at the corresponding level. By supporting progression of dystrophic spinal and osteovertebral junction disorders, chronic bronchopulmonary inflammation participates in formation of interrelated respiratory affections. New approaches to rehabilitation of CB patients with spinal lesions lie in use of pharmacopuncture, acupuncture, deep reflex-muscular massage and therapeutic exercise in consensual sequence.
HIV+ subjects have shown impairment on tests of executive function including automatic attention and verbal tasks. Impairment of semantic priming in HIV patients would suggest a disruption of automatic semantic activation. We examined semantic priming in HIV+ individuals and HIV- control participants with no history of substance abuse, neurologic or psychiatric disorder unrelated to HIV. HIV+ participants were divided into cognitively normal and cognitively impaired subgroups on the basis of a neuropsychological battery of 15 tests. Participants were presented with English words and nonword letter strings and indicated if the stimulus was a word or nonword. The nonwords were orthographically and phonologically correct and were created by rearranging the letter sequence of words ("ulpit"). All words had an obvious antonym ("deep"); two-thirds were presented as sequential antonym pairs ("enter"-"exit"). There were no group differences in speed of response to nonwords, indicating no generalized reaction time deficit. While control and cognitively normal HIV+ participants showed an effect of priming on reaction time to correctly detected words, cognitively impaired HIV+ participants did not. The lack of semantic priming demonstrated by cognitively impaired HIV+ participants suggests that they have lessened activation of automatic semantic networks.
PURPOSE: To determine the earliest changes that occur in the retina after the onset of ectopic expression of vascular endothelial growth factor (VEGF) by photoreceptors in transgenic mice, to characterize the development of neovascularization (NV), and to determine the feasibility of using these mice to test the efficacy of antiangiogenic agents. METHODS: The time course of expression of VEGF transgene mRNA was determined by reverse transcription-polymerase chain reaction (RT-PCR). Histopathologic changes in the retina were investigated by light and electron microscopy and immunocytochemistry. Standard and confocal fluorescence microscopy and image analysis were used to evaluate NV in retinal whole mounts. RESULTS: VEGF transgene mRNA was first detected in the retina by RT-PCR on postnatal day 6 (P6) and increased over the next several days to reach a constant steady-state level between P14 and P21. Abnormal cells were seen in the outer nuclear layer on P10 and among photoreceptors on P14; by P18 there were cell aggregates in the subretinal space with evidence of lumen formation. The invading cells were demonstrated to be endothelial cells by staining with an endothelial cell-specific lectin. Whole mounts of retinas perfused with fluorescein-labeled dextran showed a similar sequence of events, with sprouts from retinal vessels in the deep capillary bed seen on P14 and vessels reaching the subretinal space by P18. Confocal and standard fluorescence microscopy and changes in the number and area of neovascular lesions in the subretinal space over time measured by image analysis suggest gradual enlargement and coalescence of vascular complexes. The subretinal NV was progressively engulfed by the retinal pigmented epithelium. Invasion of blood vessels from the choroid was not identified in any specimen. CONCLUSIONS: These data support the feasibility of using rhodopsin-VEGF transgenic mice to study tissue-specific aspects of NV in the retina and to test antiangiogenic agents for inhibition of intraretinal and subretinal NV.
We examined the morphology, physiology, and 16S rRNA gene sequences of three culture collection strains and of ten novel isolates of unicellular cyanobacteria from hypersaline environments. The strains were morphologically diverse, with average cell widths ranging from 2.8 to 10.3 micron. There were single-celled, colonial, and baeocyte-forming strains. However, morphological traits were markedly variable with culture conditions. In contrast, all strains displayed extreme halotolerance (growing close to optimally at above 12% salinity); all were obligately marine, euryhaline, and moderately thermophilic; and all shared a suite of chemotaxonomic markers including phycobilins, carotenoids, and mycosporine-like amino acids. 16S rRNA gene sequence analysis indicated that the strains were related to each other. Sequence similarity analysis placed the strains in a monophyletic cluster (which we named the Halothece cluster) apart from all cultured or uncultured, not extremely halotolerant cyanobacteria whose 16S rRNA gene sequences are available in public nucleotide sequence databases. This represents the first case in which a phylogenetically coherent group of cyanobacteria can be defined on the basis of physiology. The Halothece cluster contained two subclusters that may be divergent at the generic level, one encompassing 12 strains (spanning 5% 16S rRNA gene sequence divergence and named the Euhalothece subcluster), and a single deep-branching isolate. Phenotypic characterization of the isolates, including morphological, physiological, and chemotaxonomic traits, did not distinguish these subclusters and only weakly suggested the existence of two separate clades, one encompassing strains of small cell size (cell width < 5 m) and another one encompassing strains of larger cell size.
The main object of this study was to investigate the molecular basis for changes in the spectral sensitivity of the visual pigments of deep-sea fishes. The four teleost species studied, Hoplostethus mediterraneus, Cataetyx laticeps, Gonostoma elongatum and Histiobranchus bathybius, are phylogenetically distant from each other and live at depths ranging from 500 to almost 5000 m. A single fragment of the intronless rod opsin gene was PCR-amplified from each fish and sequenced. The wavelength of peak sensitivity for the rod visual pigments of the four deep-sea species varies from 483 nm in H. mediterraneus and G. elongatum to 468 nm in C. laticeps. Six amino acids at sites on the inner face of the chromophore-binding pocket formed by the seven transmembrane a-helices are identified as candidates for spectral tuning. Substitutions at these sites involve either a change of charge, or a gain or loss of a hydroxyl group. Two of these, at positions 83 and 292, are consistently substituted in the visual pigments of all four species and are likely to be responsible for the shortwave sensitivity of the pigments. Shifts to wavelengths shorter than 480 nm may involve substitution at one or more of the remaining four sites. None of the modifications found in the derived sequences of these opsins suggest functional adaptations, such as increased content of hydroxyl-bearing or proline residues, to resist denaturation by the elevated hydrostatic pressures of the deep sea. Phylogenetic evidence for the duplication of the rod opsin gene in the Anguilliform lineage is presented.
MOTIVATION: Long non-coding RNAs (lncRNAs) have emerged as crucial players in diverse physiological and pathological processes, yet the biological mechanisms of the vast majority of lncRNAs remain elusive. To fill this gap, it is necessary to improve the accuracy of lncRNA identification and functional annotation. RESULTS: Here, we introduce LncADeep 2.0, an integrated deep learning framework designed to meet these needs. In the identification module, LncADeep 2.0 incorporated novel peptide features along with sequence and structural information, demonstrating superior performance over our previous LncADeep and other existing tools on both annotated transcripts from GENCODE and RNA-seq data. For functional annotation, LncADeep 2.0 leveraged lncRNA-centric interaction networks and gene ontology terms through the transfer learning strategy to achieve robust annotation performance with limited functional data. Compared to LncADeep, LncADeep 2.0 could accurately elucidate the general functions of given lncRNA sequences, predict tissue- or cell-type-specific functions from bulk and single-cell RNA-seq data, and establish connections between tumor-associated lncRNAs and genomic markers. Overall, LncADeep 2.0 stands out as an efficient and reliable tool for lncRNA identification and functional annotation across a wide spectrum of biological processes. AVAILABILITY AND IMPLEMENTATION: LncADeep 2.0 is available for use at https://github.com/Jefferson-Chou/LncADeep2 and https://doi.org/10.5281/zenodo.17164767.
Deep-sea soft sediments from trench systems and depths in the northwestern Pacific Ocean ranging from less than 300 to 10,897 m in depth have been analyzed for three target genera of actinomycetes: Micromonospora, Rhodococcus, and Streptomyces. Only culturable strains, recovered at atmospheric pressure on selective isolation media, have been examined to date. Maximum recoveries of culturable bacteria were greater that 10(7)/ml wet g sediment, but actinomycetes comprised a small proportion of this population (usually less than 1%). The target actinomycetes were isolated at all depths except from the Mariana Trench sediments. Actinomycete colonies were defined initially on the basis of colony morphologies, and preliminary identification then was made by chemotaxonomic tests. Pyrolysis mass spectrometry (PyMS) of deep-sea mycolic acid-containing actinomycetes gave excellent correspondence with numerical (phenetic) taxonomic analyses and subsequently was adopted as a rapid procedure for assessing taxonomic diversity. PyMS analysis enabled several clusters of deep-sea rhodococci to be distinguished that are quite distinct from all type strains. 16S rRNA gene sequence analysis has revealed that several of these marine rhodococci have sequences that are very similar to certain terrestrial species of Rhodococcus and to Dietzia. There is evidence for the intrusion of terrestrial runoff into these deep trench systems, and the inconsistency of the phenotypic and molecular taxonomies may reflect recent speciatiion events in actinomycetes under the high-pressure conditions of the deep sea. The results of DNA-DNA pairing experiments point to the novelty of Rhodococcus strains recovered from hadal depths in the Izu Bonin Trench. Biotransformation studies of deep-sea bacteria have focused on nitrile compounds. Nitrile-metabolizing bacteria, closely related to rhodococci, have been isolated that grow well at low temperature, high salt concentrations, and high pressures, suggesting that they are of marine origin or have adapted to the deep-sea environment.
The cDNAs encoding two dimeric hemoglobins, Hbs I and II, of the deep-sea clam Calyptogena soyoae were amplified by PCR and the complete nucleotide sequences determined. The cDNA-derived amino acid sequences agreed completely with those determined chemically. Many of the molluscan intracellular globin genes have a characteristic four-exon/three-intron structure, with the precoding and two conventional introns conserved widely in animal globin genes. In this work we have determined the exon/intron organization of two hemoglobin genes of the deep-sea clam C. soyoae. Surprisingly, this gene has no precoding intron but instead contains an additional intron in the A-helix (A3.1), together with the two conventional introns (B12.2 and G6.3). This observation suggests that the precoding intron has been lost and the insertion of intron in A-helix occurred in the genes of Calyptogena. Alternatively, the sliding of intron from precoding to A-helix might have occurred.
Single-gene trees have failed to locate the root of the eukaryote tree because of systematic biases in sequence evolution. Structural genetic data should yield more reliable insights into deep phylogenetic relationships. We searched major protist groups for the presence or absence of a gene fusion in order to locate the root of the eukaryote tree. In striking contrast to previous molecular studies, we show that all eukaryote groups ancestrally with two cilia (bikonts) are evolutionarily derived. The root lies between bikonts and opisthokonts (animals, Fungi, Choanozoa). Amoebozoa either diverged even earlier or are sister of bikonts or (less likely) opisthokonts.
The crystal structure of the signal sequence binding subunit of the signal recognition particle (SRP) from Thermus aquaticus reveals a deep groove bounded by a flexible loop and lined with side chains of conserved hydrophobic residues. The groove defines a flexible, hydrophobic environment that is likely to contribute to the structural plasticity necessary for SRP to bind signal sequences of different lengths and amino acid sequence. The structure also reveals a helix-turn-helix motif containing an arginine-rich alpha helix that is required for binding to SRP RNA and is implicated in forming the core of an extended RNA binding surface.
PURPOSE: Advancements in sequencing technologies have significantly improved clinical genetic testing; yet, the diagnostic yield remains around 30% to 40%. Emerging technologies are now being deployed to address the remaining diagnostic gap. METHODS: We tested whether short-read genome sequencing could increase the diagnostic yield in individuals enrolled into the UCI-GREGoR research study, who had suspected Mendelian conditions and prior inconclusive testing. Two other collaborative research cohorts, focused on aortopathy and dilated cardiomyopathy, consisted of individuals who were undiagnosed but had not undergone harmonized prior testing. RESULTS: We sequenced 353 families (754 participants) and found a molecular diagnosis in 54 (15.3%) of them. Of these diagnoses, 55.5% were previously missed because the causative variants were in regions not originally interrogated. In 5 cases, they were deep intronic variants, all of which led to abnormal splicing and pseudoexons, as directly shown by RNA sequencing. All 5 of these variants had inconclusive spliceAI scores. In 26% of newly diagnosed cases, the causal variant could have been detected by exome sequencing reanalysis. CONCLUSION: Genome sequencing can overcome limitations of clinical genetic testing, such as the inability to call intronic variants. Our findings highlight pseudoexons as a common mechanism via which deep intronic variants cause Mendelian disease.
The neighbor-joining (NJ) method is widely used in reconstructing large phylogenies because of its computational speed and the high accuracy in phylogenetic inference as revealed in computer simulation studies. However, most computer simulation studies have quantified the overall performance of the NJ method in terms of the percentage of branches inferred correctly or the percentage of replications in which the correct tree is recovered. We have examined other aspects of its performance, such as the relative efficiency in correctly reconstructing shallow (close to the external branches of the tree) and deep branches in large phylogenies; the contribution of zero-length branches to topological errors in the inferred trees; and the influence of increasing the tree size (number of sequences), evolutionary rate, and sequence length on the efficiency of the NJ method. Results show that the correct reconstruction of deep branches is no more difficult than that of shallower branches. The presence of zero-length branches in realized trees contributes significantly to the overall error observed in the NJ tree, especially in large phylogenies or slowly evolving genes. Furthermore, the tree size does not influence the efficiency of NJ in reconstructing shallow and deep branches in our simulation study, in which the evolutionary process is assumed to be homogeneous in all lineages.
Three Gram-positive bacteria designated CV-2T, CV-40 and AC-30 were isolated from a highly alkaline groundwater environment (pH 11.4). These organisms formed very small rod-shaped cells, are aerobic, non-spore-forming, catalase-positive, oxidase-negative, with an optimum growth temperature of 35 degrees C and optimum pH value of growth between 8.5 and 9.0. The strains possessed a novel B-type cell-wall peptidoglycan structure with lysine as the diamino acid; the major respiratory quinones were menaquinone 12 (MK12) and MK13. The G + C content of DNA was between 67.1 and 70.7 mol%. The phylogenetic analyses of the sequences of the 16S rRNA genes reveled that they formed a deep branch within the family Microbacteriaceae, with the highest similarity of approximately 95.6% with members of the genera Agreia, Agrococcus, Cryobacterium, Clavibacter, Frigoribacterium, Leifsonia, Mycetocola, Rhodoglobus, Salinibacterium and Subtercola. Based on the phylogenetic analyses and distinct phenotypic characteristics, we are of the opinion that strains CV-2T, CV-40 and AC-30, represent a new species of a novel genus within the family Microbacteriaceae for which we propose the name Microcella putealis gen. nov., sp. nov.
To contribute to the understanding of deep-sea planktonic communities, we explored the prokaryotic diversity of a 3000 m deep site at the Antarctic Polar Front using molecular methods. Bacterial 16S rDNA-amplified sequences corresponded to the as yet uncultivated groups SAR11, within the alpha-Proteobacteria, and SAR324, within the delta-Proteobacteria, as well as to the gamma-Proteobacteria, Cytophagales, Planctomyces, Gram-positives, and the group of environmental sequences SAR406. Among them, gamma-proteobacterial sequences were the most abundant and diverse. Within Archaea, and using six different primer sets for 16S rDNA amplification, only euryarchaeotal sequences were retrieved. Most of them clustered with the Thermoplasma-related marine groups II and III, but some corresponded to a recently described group of marine sequences emerging at the base of haloarchaea. Our data suggest that gamma-Proteobacteria and Euryarchaeota may be dominant elements in terms of genetic diversity of the two prokaryotic domains in this deep-sea pelagic area.
Standard ambulatory night sleep electroencephalograph (EEG) of 11 long-term practitioners of the Transcendental Meditation (TM) program reporting "higher states of consciousness" during sleep (the experimental group) was compared to that of nine short-term practitioners and 11 non-practitioners. EEG tracings during stages 3 and 4 sleep showed the experimental group to have: 1) theta-alpha activity simultaneously with delta activity and 2) decreased chin electromyograph (EMG) during deep sleep (p = 0.002) compared to short-term practitioners. Spectral analysis fast Fourier transform (FFT) data of the first three cycles showed that: 3) the experimental subjects had significantly greater theta 2 (6-8 Hz)-alpha 1 (8-10 Hz) relative power during stages 3 and 4 than the combined control groups [t(30) = 5.5, p = 0.0000008] with no difference in time in delta; 4) there was a graded difference across groups during stages 3 and 4 in theta 2-alpha 1 power, with experimentals having greater power than short-term practitioners, who in turn had greater power than non-practitioners [t(30) = 5.08, p = 0.00002]; and 5) experimentals also had increased rapid eye movement (REM) density during REM periods compared to short-term practitioners (p = 0.04). Previous studies have found increased theta-alpha EEG activity during reported periods of "transcendental consciousness" during the TM technique. In the Vedic tradition, as described by Maharishi Mahesh Yogi, transcendental consciousness is the first of a sequence of higher states. The maintenance of transcendental consciousness along with deep sleep is said to be a distinctive criterion of further, stabilized higher states of consciousness. The findings of this study are interpreted as physiological support for this model.
A possibility was studied of applying the deep and infrequent respiration test with an object of increasing the informativeness of the statistical parameters of the ECG sequence interval as applicable to the quantitative evaluation of the functional state of man. Examination of patients with wound infection shown that the deep respiration test makes in possible to assess quantitatively the status of test subjects on the basis of two statistical parameters (the mean value of RR interval of ECG and root-mean-square deviation).
The conger eel (Conger conger) is a nocturnal fish that can be found living in shallow coastal water and deep water down to 1000 m. The conger eel has a pure rod retina with a visual pigment maximally sensitive to blue light around 487 nm. We have cloned and sequenced the opsin cDNA which is presumed to code for this visual pigment and have found it to be highly homologous to the form of opsin that is expressed in mature, deep-living European eels (Anguilla anguilla). The opsin sequence information presented here provides additional evidence that specific amino acid sites are involved in the spectral tuning of this class of blue-sensitive visual pigments.