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Results for “spatial sequencing”

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PubMed1993

Negative supercoiling and nucleosome cores. II. The effect of negative supercoiling on the positioning of nucleosome cores in vitro.

The influence of unrestrained negative superhelical stress on nucleosome core positioning was investigated in vitro for a core located on a section of the early H1-H4 histone gene spacer of Psammechinus miliaris. We show that the position of this core on a reconstituted molecule occupied by 11 nucleosome cores is identical on a linear DNA molecule and a circular DNA molecule in the absence of unrestrained negative superhelical stress. This position is also identical to that previously found on a 337 base-pair fragment of corresponding sequence. We conclude that the core position is determined primarily by the DNA sequence, and is not influenced by core-core interactions or spatial constraints imposed by an altered geometry of the DNA molecule. This finding is supported by the identical positions assumed by the nucleosome core after altering the angular orientation of the DNA molecule, and presumably that of adjacent cores, on either one or both sides of the test core. It is further demonstrated that the core on the histone spacer region assumes identical positions on circular DNA molecules in both the presence and absence of excess negative superhelical stress equivalent to sigma = -0.03. This result indicates that conservative levels of negative supercoiling do not induce a shift in the positions of nucleosome cores. The biological implications of the experimental results are discussed and related to the findings of other workers.

Base Sequence↗

PubMed1996

Cardiac activation mapping by MRI.

To establish cardiac MRI as a tool for noninvasive evaluation of activation patterns, 10 healthy volunteers were examined by cine segmented turboFLASH imaging sequences. Sequence modifications for low signal blood-pool appearance were applied, i.e., bilateral spatial saturation for segmented turboFLASH imaging. Pixelwise calculation of first-harmonic Fourier phase values (displayed as color-encoded maps) reveal either anterior septal or left ventricular free-wall sites as areas of earliest phase spreading towards posterior paraseptal sites in segmented turboFLASH scans. Phase scatter is lower in unsaturated than spatially presaturated segmented turboFLASH studies. Phase standard deviation in areas of endocardial displacement is higher in basal than apical slice positions in these scans. Early results indicate that first-harmonic Fourier phase analysis of cardiac-segmented turboFLASH MRI cine studies may provide a tool for noninvasive studies of cardiac activation sequence.

Adult↗

PubMed1992

Distinct spatial and temporal expression patterns of K+ channel mRNAs from different subfamilies.

Different types of K+ channels play important roles in many aspects of excitability. The isolation of cDNA clones from Drosophila, Aplysia, Xenopus, and mammals points to a large multigene family with several distinct members encoding K+ channels with unique electrophysiological and pharmacological properties. Given the pivotal role K+ channels play in the fine tuning of electrical properties of excitable tissues, we studied the spatial and temporal basis of K+ channel diversity. We report the isolation of two putative K+ channels that define two new subfamilies based upon amino acid sequence similarities with other known K+ channels. Northern blot and in situ hybridization studies revealed differences in the spatial and temporal expression patterns for these two new clones along with mRNAs from other K+ channel subfamilies. Two of the K+ channels studied are predominantly expressed in the brain. One of the "brain-specific" K+ channels is first expressed after about 2 weeks of postnatal cerebellar development and remains at levels about 10-fold higher in the cerebellum than in the rest of the brain.

Amino Acid Sequence↗

PubMed1988

Imaging analysis and selected sequence monitoring of small peptides using planar chromatography/secondary ion mass spectrometry.

Mixtures of enkephalins and bradykinins contained on silica gel thin-layer chromatograms were imaged by secondary ion mass spectrometry. Spatial profiles of the sample distribution in the chromatographic plane were determined using a precision manipulator that moves the sample into and out of the primary beam generated in a liquid metal gallium source. For instance, spatial profiles are conveniently measured by monitoring the protonated molecule [M + H]+ of each compound or, in a selected-sequence monitoring experiment, by monitoring a sequence ion common to the members of each peptide group. The latter experiment is useful for quick location and identification of a class of peptides separated by planar chromatography, including thin-layer chromatography and electrophoresis.

Bradykinin↗

PubMed1990

Position-specific activity of the Hox1.1 promoter in transgenic mice.

During development, positional values have to be assigned to groups of cells. The murine Hox genes are a class of genes that are predicted to be involved at some stage in this process. During embryogenesis they are expressed in distinct overlapping region- and stage-specific patterns and therefore must be regulated in response to positional information. In this study, we have analysed the activity of Hox1.1 promoter sequences in transgenic mice. The use of lacZ as a marker allows a detailed analysis of expression at the single cell level during early embryonic development. We show that 3.6 kbp of promoter and 1.7 kbp of 3' sequences provide sufficient regulatory information to express a transgene in a spatial and temporal manner indistinguishable from the endogenous Hox1.1 gene during the period of development when Hox1.1 expression is established. The activation occurs in a strict order in specific ectodermal and mesodermal domains. Within each of these domains the transgene is activated over a period of four hours apparently randomly in single cells. In a following second period, Hox1.1 and transgene expression patterns diverge. In this period, transgene expression persists in many mesodermally derived cells that do not express Hox1.1 indicating the absence of a negative regulatory element in the transgene. The anterior boundary of transgene expression is identical to that of Hox1.1. However, no posterior boundary of transgene expression is set, suggesting that a separate element absent from the transgene specifies this boundary.

Animals↗

PubMed2006

[MRT sequences. Part I].

When magnetic resonance tomography is used in clinical practice a large number of different imaging techniques (sequences) are applied for the imaging. Despite this large number of sequences, which is often difficult to keep track of, all types of sequence work largely according to the same or very similar principles. In this continuing education course these principles are elaborated and the elements that make up a sequence are explained. These elements working together make it possible to assign the signals received from the receiver coil to their point of origin within the body. This first part of the further training is devoted largely to basic observations on spatial encoding of the signals, but also to the difference between 2D and 3D sequences. In addition, the techniques generally used in clinical practice for multislice imaging with 2D images are discussed in some detail.

Algorithms↗

PubMed2004

Spatial variation in Streptomyces genetic composition and diversity in a prairie soil.

Understanding how microbial genotypes are arrayed in space is crucial for identifying local factors that may influence the spatial distribution of genetic diversity. In this study we investigated variation in 16S rDNA sequences and rep-PCR fingerprints of Streptomyces stains isolated from prairie soil among three locations and four soil depths. Substantial variation in Streptomyces OTU (operational taxonomic unit) and BOX-PCR fingerprint diversity was found among locations within a limited spatial area (1 m2). Further, phylogenetic lineages at each location were distinct. However, there was little variation in genetic diversity among isolates from different soil depths and similar phylogenetic lineages were found at each depth. Some clones were found at a localized scale while other clones had a relatively widespread distribution. There was poor correspondence between 16S rDNA groupings and rep-PCR fingerprint groupings. The finding of distinct phylogenetic lineages and the variation in spatial distribution of clones suggests that selection pressures may vary over the soil landscape.

DNA Fingerprinting↗

PubMed1992

DNA sequence variation of the mitochondrial control region among geographically and morphologically remote European brown trout Salmo trutta populations.

Throughout its natural range, the brown trout Salmo trutta L. exhibits a complex pattern of morphological and life-history variation. This has led to considerable taxonomic confusion, hampering the understanding of the evolutionary history of the species. To document the phylogenetic relationships among morphologically and geographically remote brown trout populations across western Europe, we determined the DNA sequence variation in segments of the mitochondrial control region for 151 individuals representing 24 populations. DNA was prepared for double-stranded sequencing by the polymerase chain reaction (PCR). Twenty-one variable nucleotide positions within a 640-bp fragment surveyed defined 12 genotypes differing by a mean of 7 nucleotide substitutions (range 1-12). Five major phylogenetic assemblages differing by mean sequence divergence estimates of 0.96 to 1.44% were identified. These groupings exhibited a strong spatial partitioning but lacked congruence with either ecological or morphological differentiation. Complete mitochondrial DNA (mtDNA) monomorphism across all Atlantic basin populations contrasted with the high interdrainage genetic diversity observed in more southerly populations. This study exemplified the usefulness of mitochondrial DNA sequence analysis for estimating phylogenetic relationships within S. trutta populations.

Animals↗

PubMed2001

The effects of lesions to the rat hippocampus or rhinal cortex on olfactory and spatial memory: retrograde and anterograde findings.

The role of the hippocampal system in retrograde and anterograde amnesia was investigated by using a novel olfactory-guided paradigm and a traditional test of spatial learning. In the retrograde study, rats were trained on a sequence of two-choice olfactory discriminations in the weeks prior to receiving neurotoxic lesions of the hippocampus or aspiration lesions of the perirhinal-entorhinal cortex. Memory tests for preoperatively learned discriminations revealed no statistical impairment for subjects with damage to the hippocampus on a problem learned remote in time from surgery (i.e., 4 weeks +) or on the two recently learned discriminations (i.e., 1-3 weeks prior to surgery). The performance of subjects with perirhinal-entorhinal damage provided an important comparison for subjects with specific hippocampal lesions. Despite showing intact memory for the remotely learned problem, perirhinal-entorhinal damage resulted in numerically (although not significantly) weaker performance on postoperative tests of retention for the discriminations learned in the 3 weeks prior to surgery. In the anterograde portion of the study, long-term memory for newly acquired discriminations was spared in subjects with damage to the hippocampus, whereas subjects in the perirhinal-entorhinal lesion group again showed the weakest memory performance on these tests of 5-day retention. Postoperative water maze learning was uniformly impaired in subjects with damage to the hippocampus and perirhinal-entorhinal cortex, thus confirming the effect of these lesions and supporting the involvement of these brain areas in spatial processes. These findings further dissociate the specific involvement of the hippocampus in tasks of a spatial-relational nature versus nonrelational tasks, such as discrimination learning and recognition memory (e.g., Duva et al., 1997; Eichenbaum, 1997; Eichenbaum, Schoenbaum, Young, & Bunsey, 1996). Moreover, the results suggest that damage to the hippocampus itself does not contribute to retrograde or anterograde memory impairments for all types of information, whereas the data suggest a more important role for the perirhinal-entorhinal cortex in recognition memory, irrespective of modality.

Animals↗

PubMed2025

Multi-omics analysis reveals distinct spatial compartmentalization of lung repair niches in pediatric ARDS.

BACKGROUND: Pediatric acute respiratory distress syndrome (PARDS), often triggered by viral infections, is a life-threatening condition. Despite its severity, children demonstrate significantly better survival rates and superior lung repair compared to adults. However, the mechanisms underlying this age-specific advantage remain incompletely understood. PATIENTS AND METHODS: We conducted a pilot multi-omics study of influenza-associated PARDS integrating single-cell RNA sequencing (scRNA-seq) of pediatric lung tissue and bronchoalveolar lavage fluid (BALF), spatial transcriptomics, and plasma proteomics. Analyses were harmonized with the Human Lung Cell Atlas (HLCA) reference, reanalysis of public pediatric PARDS airway scRNA-seq, and contextual comparisons to adult lethal COVID-19 lung. RESULTS: Tissue scRNA-seq and spatial data indicated outcome-linked divergence in PARDS. Survivor showed spatially restricted repair with preserved alveolar type II (AT2) cells, AT2-to-alveolar type I (AT1) differentiation signatures, and higher KRT17, whereas fatal case and adults exhibited diffuse immune activation with pro-fibrotic and pro-apoptotic signaling. In BALF, KRT17-positive airway stress–repair epithelial cells (hillock-like) increased from the acute to recovery phase, and plasma proteomics showed higher circulating KRT17 in survivors. HLCA-based label transfer strengthened cell-type definitions and enabled pediatric–adult comparisons suggesting biological and developmental differences; the adult lethal COVID-19 atlas provided a benchmark with attenuated epithelial repair and prominent collagen CTHRC1-pathologic fibroblasts. Fibroblast programs were regionally compartmentalized, with injury-enriched CTHRC1+ states versus alveolar fibroblasts in preserved areas, and showed stronger injury–homeostasis anti-correlation in fatalities. Myeloid remodeling included BALF transitions from FCN1-high inflammatory states toward FABP4-positive resident-like states, consistent with public pediatric datasets showing reduced inflammatory and interferon-stimulated gene (ISG) modules and severity-linked increases in aged neutrophils. CONCLUSIONS: This pilot multi-omics case series outlines putative pediatric lung repair niches in influenza-associated PARDS. KRT17-positive transitional epithelium, preserved AT2 differentiation, and restoration of resident-like macrophages may align with recovery, whereas diffuse immune activation and CTHRC1-enriched fibroblast programs may accompany worse outcomes. HLCA-guided annotations and adult benchmarks indicate possible age-related differences, warranting validation in larger multi-center cohorts.

Humans↗

PubMed2005

Molecular characterization and expression of the UV opsin in bumblebees: three ommatidial subtypes in the retina and a new photoreceptor organ in the lamina.

Ultraviolet-sensitive photoreceptors have been shown to be important for a variety of visual tasks performed by bees, such as orientation, color and polarization vision, yet little is known about their spatial distribution in the compound eye or optic lobe. We cloned and sequenced a UV opsin mRNA transcript from Bombus impatiens head-specific cDNA and, using western blot analysis, detected an eye protein band of approximately 41 kDa, corresponding to the predicted molecular mass of the encoded opsin. We then characterized UV opsin expression in the retina, ocelli and brain using immunocytochemistry. In the main retina, we found three different ommatidial types with respect to the number of UV opsin-expressing photoreceptor cells, namely ommatidia containing two, one or no UV opsin-immunoreactive cells. We also observed UV opsin expression in the ocelli. These results indicate that the cloned opsin probably encodes the P350 nm pigment, which was previously characterized by physiological recordings. Surprisingly, in addition to expression in the retina and ocelli, we found opsin expression in different parts of the brain. UV opsin immunoreactivity was detected in the proximal rim of the lamina adjacent to the first optic chiasm, which is where studies in other insects have found expression of proteins involved in the circadian clock, period and cryptochrome. We also found UV opsin immunoreactivity in the core region of the antennal lobe glomeruli and different clusters of perikarya within the protocerebrum, indicating a putative function of these brain regions, together with the lamina organ, in the entrainment of circadian rhythms. In order to test for a possible overlap of clock protein and UV opsin spatial expression, we also examined the expression of the period protein in these regions.

Amino Acid Sequence↗

PubMed2006

DNA sequence-directed assembly of two peptide bioconjugates.

Peptide bioconjugates combine the molecular recognition features of the polypeptide from a DNA-binding protein and the DNA-sensitive fluorescence of an intercalating dye. Here, DNA template-directed assembly of two bioconjugate probes was examined by steady-state fluorescence resonance energy transfer and time-resolved single photon counting. The Förster critical distance was determined to be approximately 26 A for the oxazole yellow and thiazole orange donor-acceptor pair. The efficiency of energy transfer for two bioconjugates was a function of the number of intervening base pairs between two DNA cognate sites. These probes were sufficiently sensitive to detect sequence dependent curvature and polypeptide induced bending of the DNA. Molecular probes capable of examining spatial aspects of protein complexes at promoter sites could yield important information about the early events in transcription initiation.

Amino Acid Sequence↗

PubMed1997

Pre-processing variance reducing techniques in multispectral positron emission tomography.

Stochastic fluctuations and systematic errors severely restrict the potential of multispectral acquisition to improve scatter correction by energy-dependent processing in high-resolution positron emission tomography (PET). To overcome this limitation, three pre-processing approaches which reduce stochastic fluctuations and systematic errors without degrading spatial resolution were investigated: statistical variance was reduced by smoothing acquired data in energy space, systematic errors due to nonuniform detector efficiency were minimized by normalizing the data in the spatial domain and the overall variance was further reduced by selecting an optimal pre-processing sequence. Selection of the best protocol to reduce stochastic fluctuations entailed comparisons between four smoothing algorithms (prior constrained (PC) smoothing, weighted smoothing (WS), ideal low-pass filtering (ILF) and mean median (MM) smoothing) and permutations of three pre-processing procedures (smoothing, normalization and subtraction of random events). Results demonstrated that spectral smoothing by WS, ILF and MM efficiently reduces the statistical variance in both the energy and spatial domains without observable spatial resolution loss. The ILF algorithm was found to be the most convenient in terms of simplicity and efficiency. Regardless of the position of subtraction of randoms in the sequence, reduction of the systematic errors by normalization followed by spectral smoothing to suppress statistical noise produced the best results. However, subtraction of random events first in the sequence reduces computation load by half since the need to pre-process this distribution before subtraction is removed. In summary, normalizing data in the spatial domain and smoothing data in energy space are essential steps required to reduce systematic errors and statistical variance independently without degrading spatial resolution of multispectral PET data.

Biophysical Phenomena↗

PubMed1996

Spatial and temporal variations in extracellular matrix of periocular and corneal regions during corneal stromal development.

The development of the avian corneal stroma occurs in discrete developmental stages. During this sequence of events, the neural crest-derived corneal fibroblast precursor cells are surrounded by distinct extracellular matrices which change both spatially and temporally. To elucidate the role of these matrices, extracellular matrix components in the periocular mesenchyme and cornea were analysed prior to and during migration and differentiation of corneal fibroblasts using antibodies against collagens, proteoglycans and glycoproteins. Previous work has concentrated on the matrix of the corneal stroma rather than the matrix of the periocular mesenchyme. Since the precursors of the corneal fibroblasts are present within the must migrate through the periocular mesenchyme prior to entry into the cornea proper, this environment was fully evaluated. The present study documents the matrix composition of both the cornea and periocular mesenchyme at developmental stages that are prior to and after initiation of corneal invasion by the corneal fibroblast precursors. Variations in matrix molecules comprising both the periocular mesenchyme and cornea were demonstrated. These include changes in the distribution of collagen types I, II, III, IV and VI; the proteoglycans decorin and lumican; as well as the adhesive glycoproteins tenascin, fibronectin and laminin. It is hypothesized that the variations in matrix localization are important in the regulation of cell migration and differentiation during normal corneal development. Any regulation is likely to involve a combination of components found in the extracellular matrices and therefore, a consideration of the matrix rather than isolated components is required.

Animals↗

PubMed2001

Lysyl oxidases: a novel multifunctional amine oxidase family.

Lysyl oxidase (LOX), a copper-containing amine oxidase, belongs to a heterogeneous family of enzymes that oxidize primary amine substrates to reactive aldehydes. LOX has been traditionally known for one function, the extracellular catalysis of lysine-derived cross-links in fibrillar collagens and elastin. More recently, diverse roles have been attributed to lysyl oxidase and these novel activities cover a spectrum of diverse biological functions such as developmental regulation, tumor suppression, cell motility, and cellular senescence. Lysyl oxidase has also been shown to have both intracellular and intranuclear locations. The multifunctional properties of lysyl oxidase (LOX) and our recent discovery of three novel members of this amine oxidase family, LOX-like (LOXL), LOXL2, and LOXL3, indicate the possibility that these varied functions are performed in both intracellular and extracellular environments by individual novel members of the LOX amine-oxidase family. Structural similarities of the highly conserved copper-binding and lysyl-tyrosylquinone cofactor sites among the LOX and LOX-like proteins may result in similar amine oxidase activities. However, specific novel functions, such as a potential role in cell adhesion and cell growth control, will be determined by other, conserved domains such as the cytokine receptor-like domain that is shared by all LOXs and by multiple scavenger receptor cysteine-rich (SRCR) domains present in LOXL2 and LOXL3. Furthermore, these functions may be carried out in a temporally and spatially regulated fashion.

Amino Acid Sequence↗

PubMed2006

The functional and developmental organization of cognitive developmental sequences.

This study examines the organization and development of 5 domains of reasoning (categorical, quantitative, spatial, causal, and propositional) and the construct validity of a test designed to measure development from early adolescence to early adulthood. The theory underlying the test is first summarized and the conceptual design of the test is then illustrated. Each domain was addressed by tasks tapping abilities known to be acquired in this age period. The test was administered to 629 adolescents ranging in age from 12 to 18 years. Confirmatory factor analysis validated the 5 domains of reasoning and revealed a common factor underlying all domains. The Rasch model was used to scale the items and specify the reliability of the test across the whole sample and within different groups of participants (female, male, students of gymnasium, and students of lyceum). This model showed that the test is highly reliable and invariant across groups. Cluster analysis and the saltus model were applied to uncover successive developmental stage-like levels of difficulty and showed the presence of five such levels. The procedural and representational characteristics of these levels were also specified and their implications for developmental and cognitive theory were discussed.

Adolescent↗

PubMed2005

Design criteria for engineering inorganic material-specific peptides.

Development of a fundamental understanding of how peptides specifically interact with inorganic material surfaces is crucial to furthering many applications in the field of nanobiotechnology. Herein, we report systematic study of peptide sequence-activity relationships for binding to II-VI semiconductors (CdS, CdSe, ZnS, ZnSe) and Au using a yeast surface display system, and we define criteria for tuning peptide affinity and specificity for these material surfaces. First, homohexapeptides of the 20 naturally occurring amino acids were engineered, expressed on yeast surface, and assayed for the ability to bind each material surface in order to define functional groups sufficient for binding. Histidine (H6) was able to mediate binding of yeast to the five materials studied, while tryptophan (W6), cysteine (C6), and methionine (M6) exhibited different levels of binding to single-crystalline ZnS and ZnSe and polycrystalline Au surfaces. The ability of neighboring amino acids to up- and down-modulate histidine binding was then evaluated by use of interdigitated peptides (XHXHXHX). While the 20 amino acids exhibited a unique fingerprint of modulation for each material, some general trends emerged. With neutral defined by alanine, up-modulation occurred with glycine, basic amino acids, and the previously defined binding amino acids histidine, tryptophan, cysteine, and methionine, and down-modulation generally occurred with acidic, polar, and hydrophobic residues. We conclude that certain amino acids directly bind the material surface while neighboring amino acids locally modulate the binding environment for the materials we studied. Therefore, by the specific placement of up- and down-modulating amino acids, material specificity can be controlled. Finally, by employing the compositional and spatial criteria developed herein, it was possible to predictively design peptide sequences with material specificity, including a multimaterial binder, a Au-specific binder, and a ZnS-specific binder, that were verified as such in the context of yeast display.

Amino Acid Sequence↗

PubMed2002

Myosin heavy chain expression in cranial, pectoral fin, and tail muscle regions of zebrafish embryos.

To investigate whether different myosin heavy chain (MHC) isoforms may constitute myofibrils in the trunk and tail musculature and if their respective expression may be regulated by spadetail (spt) and no tail (brachyury), we identified and characterized mRNA expression patterns of an embryonic- and tail muscle-specific MHC gene (named myhz2) during zebrafish development in wild type, spt, and ntl mutant embryos. The identified myhz2 MHC gene encodes a polypeptide containing 1,935 amino acids. Deduced amino acid comparisons showed that myhz2 MHC shared 92.6% sequence identity with that of carp fast skeletal MHC. Temporal and spatial myhz2 MHC mRNA expression patterns were analyzed by quantitative RT-PCR and whole-mount in situ hybridization using primer pairs and probes designed from the 3'-untranslated region (UTR). Temporally myhz2 MHC mRNA appears in pharyngula embryos and peaks in protruding-mouth larvae. The expression level decreased in 7-day-old hatching larvae, and mRNA expression was not detectable in adult fish. Spatially in pharyngula embryos, mRNA was localized only in the tail somite region, while in long-pec embryos, transcripts were also expressed in the two cranial muscle elements of the adductor mandibulae and medial rectus, as well as in pectoral fin muscles and the tail muscle region. Myhz2 MHC mRNA was expressed in most cranial muscle elements, pectoral fin muscles, and the tail muscle region of 3-day-old hatching larvae. In contrast, no expression of myhz2 MHC mRNA could be observed in spt prim-15 mutant embryos. In spt long-pec mutant embryos, transcripts were expressed in two cranial muscle elements and the tail muscle region, but not in pectoral fin muscles, while only trace amounts of myhz2 MHC mRNA were expressed in the remaining tail muscle region of 38 hpf and long-pec ntl mutant embryos.

Amino Acid Sequence↗