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At least 631 records · Page 35Linked to original sources
PubMed2006

HEF1-aurora A interactions: points of dialog between the cell cycle and cell attachment signaling networks.

Regulated timing of cell division cycles, and geometrical precision in the planar orientation of cell division, are critical during organismal development and remain important for the maintenance of polarized structures in adults. Mounting evidence suggests that these processes are coordinated at the centrosome through the action of proteins that mediate both cell cycle and cell attachment. Our recent work identifying HEF1 as an activator of the Aurora A kinase suggests a novel hub for such integrated signaling. We suggest that defects in components of the machinery specifying the temporal and spatial integration of cell division may induce cancer and other diseases through pleiotropic effects on cell migration, proliferation, apoptosis, and genomic stability.

Adaptor Proteins, Signal Transducing↗
PubMed2002

Historic and recent events contribute to the disease dynamics of Beijing-like Mycobacterium tuberculosis isolates in a high incidence region.

SETTING: A retrospective study in an urban setting with a high tuberculosis incidence. OBJECTIVE: To study the molecular epidemiology and disease dynamics of a prevalent Mycobacterium tuberculosis strain family, F29, a subset of the Beijing strains. DESIGN: Multi-locus DNA fingerprinting and genomic analysis was used to characterise F29 isolates. Demographic and molecular epidemiologic data were correlated with those of other isolates from the setting. The geographic locations of five F29 strain clusters were plotted using a geographic information system (GIS) and an index for geographic distribution was calculated. Their cumulative temporal occurrence was also plotted. RESULTS: The genomic similarity of F29 to the Beijing-strains was confirmed. A high degree of clustering predicted high disease transmission. Spatial distribution was mostly widespread except for one cluster. Smaller foci of transmission were observed. The temporal spread showed ongoing transmission. CONCLUSION: F29 belongs to the Beijing strain group. The prevalence and high degree of strain clustering, with limited geographic clustering, indicates that F29 strains are endemic in the study community. However, recent epidemiologic events also contributed to the disease spread. The combination of molecular epidemiologic, spatial and temporal data has enhanced our understanding of the disease dynamics of Beijing strains in our study community.

Adolescent↗
PubMed2002

Ability of the hydrophobic FGF and basic TAT peptides to promote cellular uptake of recombinant Cre recombinase: a tool for efficient genetic engineering of mammalian genomes.

Conditional mutagenesis is a powerful tool to analyze gene functions in mammalian cells. The site-specific recombinase Cre can be used to recombine loxP-modified alleles under temporal and spatial control. However, the efficient delivery of biologically active Cre recombinase to living cells represents a limiting factor. In this study we compared the potential of a hydrophobic peptide modified from Kaposi fibroblast growth factor with a basic peptide derived from HIV-TAT to promote cellular uptake of recombinant Cre. We present the production and characterization of a Cre protein that enters mammalian cells and subsequently performs recombination with high efficiency in a time- and concentration-dependent manner. Histidine-tagged Cre recombinase transduced inefficiently unless fused to a nuclear localization signal (NLS). Fusion of NLS-Cre to the fibroblast growth factor transduction peptide did not improve the transducibility, whereas fusion with the TAT peptide significantly enhanced cellular uptake and subsequent recombination. More than 95% recombination efficiency in fibroblast cells, as well as murine embryonic stem cells, was achieved after His-TAT-NLS-Cre transduction. Efficient recombination could also be obtained in primary splenocytes ex vivo. We expect that application of His-TAT-NLS-Cre, which can be produced readily in large quantities from a bacterial source, will expand our abilities to manipulate mammalian genomes.

Cells, Cultured↗
PubMed2004

cis-Regulatory activity of randomly chosen genomic fragments from the sea urchin.

In order to determine the frequency and variety of cis-regulatory elements that function during embryonic development of Strongylocentrotus purpuratus, we constructed a GFP expression vector in which to test the activity of randomly chosen genomic DNA fragments that includes a promiscuous basal promoter from the endo16 gene. This vector was demonstrated to serve as a cis-regulatory element trap. We used it to carry out an initial test for the occurrence of elements that would promote GFP expression in this genome. In the screen reported here 108 different randomly chosen DNA fragments (av. 3.8 kb) were inserted in the vector, and each was injected into > 200 zygotes. Surprisingly, 13% of the fragments tested yielded detectable levels of GFP expression in the recipient embryos. Specific patterns observed included expression in endoderm, in aboral ectoderm, and in pigment cells. The majority of active constructs expressed GFP in all spatial domains of the embryo. Elements with detectable cis-regulatory activity in the embryo occur in the sample screened, on the average, about every 30 kb, and the genome must include many thousands of such elements. On further analysis one isolate was shown to contain a gut specific element as well as one that controls expression in the secondary mesenchyme cells.

Animals↗
PubMed2003

Cell shape, division and development: the 2002 American Society for Microbiology (ASM) conference on prokaryotic development.

In the last decade, the use of cytological techniques, together with the analysis of complete genomes, has dramatically advanced our understanding of bacterial development. Work on several well-developed model systems such as Bacillus subtilis, Caulobacter crescentus, Myxococcus xanthus and Streptomyces spp., has provided us with an in-depth understanding of processes such as sporulation, multicellular behaviour and the bacterial cell cycle. At the same time, these studies have revolutionized our view of the bacterial cell and shown it to be a highly complex entity with spatial and temporal organization. The recent American Society for Microbiology (ASM) conference on prokaryotic development demonstrated that several laboratories have now started to connect data obtained through functional genomic analysis with subcellular organization, thereby generating three-dimensional regulatory networks. This meeting report highlights new findings in the field, such as regulation of protein localization during sporulation and the cell cycle, control of cell-cell interaction and the initiation of cell division.

Bacillus subtilis↗
PubMed2026

Representation learning for multi-modal spatially resolved transcriptomics data.

MOTIVATION: Spatial transcriptomics enables in-depth molecular characterization of samples on a morphology and RNA level while preserving spatial location. Integrating the resulting multi-modal data is an unsolved problem, and developing new solutions in precision medicine depends on improved methodologies. RESULTS: We introduce AESTETIK, a convolutional deep learning model that jointly integrates spatial, transcriptomics, and morphology information to learn accurate spot representations. AESTETIK yielded substantially improved cluster assignments on widely adopted technology platforms (e.g. 10x Genomics™, NanoString™) across multiple datasets. We achieved performance enhancement on structured tissues (e.g. brain) with a 21% increase in median ARI over previous state-of-the-art methods. Notably, AESTETIK also demonstrated superior performance on cancer tissues with heterogeneous cell populations, showing a 2-fold increase in breast cancer, 79% in melanoma, and 21% in liver cancer. We expect that these advances will enable a multi-modal understanding of key biological processes. AVAILABILITY AND IMPLEMENTATION: AESTETIK is implemented in Python 3 and is available as open source software at http://www.github.com/ratschlab/aestetik. The Snakemake pipeline for reproducing the results is available at http://www.github.com/ratschlab/st-rep.

Spatial Transcriptomics↗
PubMed2003

Reconstitution of gene expression from a regulatory-protein-deficient hepatitis B virus genome by cell-permeable HBx protein.

Various functions are ascribed to the HBx regulatory protein of the hepatitis B virus (HBV). Due to the low expression level of HBx, it has been difficult to correlate spatial and temporal HBx expression levels with specific functions. Based on a novel cell-permeable peptide, known as the translocation motif (TLM), cell-permeable HBx fusion proteins were generated. The TLM-HBx fusion protein is rapidly internalized from the medium into almost all cells, whereas no significant internalization was seen with wild-type HBx. The major fraction of internalized HBx protein moves from the cytoplasm to the nucleus. The cytosolic fraction, however, activates c-RAF1/extracellular-signal-related kinase 2 signalling and causes activation of activator protein 1 (AP1) and nuclear factor-kappaB. The TLM-HBx protein rescues HBV gene expression from an activator-deficient HBV genome. These results indicate that cell-permeable regulatory proteins provide a novel, efficient tool for a clearly defined, dose-dependent analysis of regulatory protein function, without affecting the integrity of the cell, and can be used for the safe reconstitution of virus production from a regulatory-protein-deficient virus genome.

Cell Line, Tumor↗
PubMed1997

Transgenic animal models: new avenues in cardiovascular physiology.

Application of molecular genetic tools to inherited cardiovascular disorders has provided important insights into the molecular mechanisms underlying cardiomyopathies, arrhythmias, blood pressure regulation, and atherosclerosis. In addition, alteration of gene expression has been observed under common cardiovascular conditions such as cardiac hypertrophy and heart failure. Recent advances in transgenic and gene-targeting approaches allow a sophisticated manipulation of the mouse genome by gene addition, gene deletion, or gene modifications. These transgenic models enable the dissection of in vivo pathways responsible for these complex disease phenotypes. This review describes tissue-specific promoters suitable for targeting candidate genes to the cardiovascular system as well as a number of valuable transgenic animal models of blood pressure regulation, atherogenesis, defects in the coagulation system, cardiac hypertrophy, myocarditis, cardiomyopathies, and heart failure. Limitations and difficulties associated with these transgenic approaches are discussed. Animal models which may provide a basis for future gene therapy of cardiovascular diseases are introduced. Finally, methods are described to regulate the spatial and temporal expression level of a transgene, to inactivate a target gene in a tissue-specific manner, and to introduce specific mutations into the genome. These recent advances in transgenic technology are expected to have a considerable impact on cardiovascular research in the near future.

Animals↗
PubMed2001

The functional genomic response of developing embryonic submandibular glands to NF-kappa B inhibition.

BACKGROUND: The proper balance between epithelial cell proliferation, quiescence, and apoptosis during development is mediated by the specific temporal and spatial appearance of transcription factors, growth factors, cytokines, caspases, etc. Since our prior studies suggest the importance of transcription factor NF-kappaB during embryonic submandibular salivary gland (SMG) development, we attempted to delineate the emergent dynamics of a cognate signaling network by studying the molecular patterns and phenotypic outcomes of interrupted NF-kappaB signaling in embryonic SMG explants. RESULTS: SN50-mediated inhibition of NF-kappaB nuclear translocation in E15 SMG explants cultured for 2 days results in a highly significant increase in apoptosis and decrease in cell proliferation. Probabilistic Neural Network (PNN) analyses of transcriptomic and proteomic assays identify specific transcripts and proteins with altered expression that best discriminate control from SN50-treated SMGs. These include PCNA, GR, BMP1, BMP3b, Chk1, Caspase 6, E2F1, c-Raf, ERK1/2 and JNK-1, as well as several others of lesser importance. Increased expression of signaling pathway components is not necessarily probative of pathway activity; however, as confirmation we found a significant increase in activated (phosphorylated/cleaved) ERK 1/2, Caspase 3, and PARP in SN50-treated explants. This increased activity of proapoptotic (caspase3/PARP) and compensatory antiapoptotic (ERK1/2) pathways is consistent with the dramatic cell death seen in SN50-treated SMGs. CONCLUSIONS: Our morphological and functional genomic analyses indicate that the primary and secondary effects of NF-kappaB-mediated transcription are critical to embryonic SMG developmental homeostasis. Relative to understanding complex genetic networks and organogenesis, our results illustrate the importance of evaluating the gene, protein, and activated protein expression of multiple components from multiple pathways within broad functional categories.

Animals↗
PubMed2001

PML bodies associate specifically with the MHC gene cluster in interphase nuclei.

Promyelocytic leukemia (PML) bodies are nuclear multi-protein domains. The observations that viruses transcribe their genomes adjacent to PML bodies and that nascent RNA accumulates at their periphery suggest that PML bodies function in transcription. We have used immuno-FISH in primary human fibroblasts to determine the 3D spatial organisation of gene-rich and gene-poor chromosomal regions relative to PML bodies. We find a highly non-random association of the gene-rich major histocompatibilty complex (MHC) on chromosome 6 with PML bodies. This association is specific for the centromeric end of the MHC and extends over a genomic region of at least 1.6 megabases. We also show that PML association is maintained when a subsection of this region is integrated into another chromosomal location. This is the first demonstration that PML bodies have specific chromosomal associations and supports a model for PML bodies as part of a functional nuclear compartment.

Cell Nucleus↗
PubMed2005

[Epigenetics as an aspect of post-genomics].

A cell transmits to its progeny the activity level of many of its genes, not just their sequence. Just like the sequence may vary through a mutation, the gene activity level may change through an "epimutation" (an epigenetic modification) which is heritable and does not entail any concomitant genetic alteration. An epimutation can have important phenotypic consequences, that eventually survive to the loss of the environmental conditions that triggered it. For instance, epimutations are responsible for the divergence between a neuron and an epithelial cell that both come from the same egg and contain the same genome complement. This phenotypic difference is much larger than the one between the neurons from two animal species with dissimilar genotypes, thereby underlining the importance of epimutations. Tradition opposes the genetic and epigenetic visions, the latter being often adequated to the DNA methylation phenomenon. However, epimutations display a rich spectrum of modes that can all fit in a unique reference system based on correlated chemical, spatial and temporal scales. This reference system allows the integration of purely genetic mutations at one of its ends, thus paving the way to a new, gradual vision that encompasses the genome and the epigenome. At the other end can be found two types of epimutations that are both wide-ranging in space and rapid in producing phenotypic alterations. Firstly, long-range rearrangements of the three-dimensional structure of the chromosome may influence gene expression in an heritable fashion. Such rearrangements seem to result from the collective dynamics of DNA-related activities, particularly transcription. Lastly, heritable regulatory states, e.g. a differentiated state that results from tipping a regulatory "toggle switch", involve components that are distributed throughout the nucleus or the cytoplasm, and possibly all the way to cell confines.

Epigenesis, Genetic↗
PubMed2026

dcHiChIP: a comprehensive Nextflow-based pipeline for multiscale analysis of chromatin architecture from HiChIP data.

MOTIVATION: Despite the growing use of HiChIP to investigate protein-directed chromatin architecture, a comprehensive and reproducible pipeline for analysing these datasets-from raw reads to multiscale 3D genome features-remains lacking. Existing tools often focus on isolated components, such as loop calling or matrix generation, but fall short in integrating structural annotation, functional enrichment, and spatial modeling within a unified framework. To address this gap, we developed dcHiChIP, a modular, scalable Nextflow-based workflow that streamlines the analysis of HiChIP data, enabling both routine processing and in-depth exploration of chromatin organization and regulatory interactions. RESULTS: dcHiChIP enables robust and reproducible analysis of HiChIP datasets across multiple scales of chromatin architecture. It accepts raw sequencing data as input and generates high-quality loop calls, domain annotations, and 3D genome models. It also performs functional annotation and motif enrichment analyses. Applied to benchmark CTCF HiChIP datasets, dcHiChIP identifies major chromatin architectural features such as TADs/CCDs, A/B compartments, and chromatin stripes, and offers efficient, end-to-end execution with support for batch processing and workflow resumability. AVAILABILITY: dcHiChIP is publicly available on GitHub at https://github.com/SFGLab/dcHiChIP, with documentation at https://sfglab.github.io/dcHiChIP/. The software version used in this study is archived at Zenodo: https://doi.org/10.5281/zenodo.22030542.

Chromatin↗
PubMed1996

Bombyxin-related peptides: cDNA structure and expression in the brain of the hornworm Agrius convolvuli [corrected].

We have cloned three cDNAs from the sweet potato hornworm Agrius convolvuli that encode precursor molecules for peptides structurally related to bombyxin, an insulin-related brain secretory peptide in Bombyx mori. The Agrius bombyxin-related peptide (ABRP) cDNAs are classified into type A and B according to their sequence similarity. The prepro-ABRPs deduced from the cDNA sequences have the insulin-like domain organization of signal peptide/B chain/C peptide/A chain. The ABRP transcripts in Agrius brain were shown to locate in four pairs of medial neurosecretory cells, the homologous group of neurosecretory cells that produce bombyxins in Bombyx brain. Genomic Southern analysis indicated the presence of multiple copies of ABRP gene in the Agrius genome. Results showed that the ABRP genes are remarkably different from the vertebrate insulin genes in the number of copy and spatial localization of the transcripts.

Amino Acid Sequence↗
PubMed2002

When reverse genetics meets physiology: the use of site-specific recombinases in mice.

The use of site-specific recombinases enables the precise introduction of defined genetic mutations into the mouse genome. In theory, any deletion, point mutation, inversion or translocation can be modeled in mice. Because gene targeting is controlled both spatially and temporally, the function of a given gene can be studied in the desired cell types and at a specific time point. This 'genetic dissection' allows to define gene function in development, physiology or behavior. In this review, we focus on the technical possibilities of Cre and other site-specific recombinases but also discuss their limitations.

Animals↗
PubMed2006

In vivo enhancer analysis of human conserved non-coding sequences.

Identifying the sequences that direct the spatial and temporal expression of genes and defining their function in vivo remains a significant challenge in the annotation of vertebrate genomes. One major obstacle is the lack of experimentally validated training sets. In this study, we made use of extreme evolutionary sequence conservation as a filter to identify putative gene regulatory elements, and characterized the in vivo enhancer activity of a large group of non-coding elements in the human genome that are conserved in human-pufferfish, Takifugu (Fugu) rubripes, or ultraconserved in human-mouse-rat. We tested 167 of these extremely conserved sequences in a transgenic mouse enhancer assay. Here we report that 45% of these sequences functioned reproducibly as tissue-specific enhancers of gene expression at embryonic day 11.5. While directing expression in a broad range of anatomical structures in the embryo, the majority of the 75 enhancers directed expression to various regions of the developing nervous system. We identified sequence signatures enriched in a subset of these elements that targeted forebrain expression, and used these features to rank all approximately 3,100 non-coding elements in the human genome that are conserved between human and Fugu. The testing of the top predictions in transgenic mice resulted in a threefold enrichment for sequences with forebrain enhancer activity. These data dramatically expand the catalogue of human gene enhancers that have been characterized in vivo, and illustrate the utility of such training sets for a variety of biological applications, including decoding the regulatory vocabulary of the human genome.

Animals↗
PubMed2005

Estimating the correlation of pairwise relatedness along chromosomes.

The 'spatial' pattern of the correlation of pairwise relatedness among loci within a chromosome is an important aspect for an insight into genomic evolution in natural populations. In this article, a statistical genetic method is presented for estimating the correlation of pairwise relatedness among linked loci. The probabilities of identity-in-state (IIS) are related to the probabilities of identity-by-descent (IBS) for the two- and three-loci cases. By decomposing the joint probabilities of two- or three-loci IBD, the probability of pairwise relatedness at a single locus and its correlation among linked loci can be simultaneously estimated. To provide effective statistical methods for estimation, weighted least square (LS) and maximum likelihood (ML) methods are evaluated through extensive Monte Carlo simulations. Results show that the ML method gives a better performance than the weighted LS method with haploid genotypic data. However, there are no significant differences between the two methods when two- or three-loci diploid genotypic data are employed. Compared with the optimal size for haploid genotypic data, a smaller optimal sample size is predicted with diploid genotypic data.

Animals↗
PubMed1999

Amount and organization of the heterochromatin in Olea europaea and related species.

The amount and spatial organization of the heterochromatin in nuclei of the shoot meristem and the frequency in the nuclear DNA of sequences belonging to a family of tandem repeats were investigated in cultivars of Olea europaea and related species. Significant differences between Olea species and between cultivars of O. europaea were observed: (i) in the spatial organization of the heterochromatin in interphase nuclei as determined by the number and surface area of the chromocentres; (ii) in genome size; and (iii) in the amount of condensed chromatin as measured by cytophotometry carried out at different thresholds of optical density. DNA elements belonging to a family of tandem repeats about 80 bp in length (OeTaq80 repeats) were isolated from the genomic DNA of an olive cultivar. It was shown: (i) by nucleotide sequence comparisons, that these repeats display variability in structure even within the same array, where different elements may share no more than 74% homology; (ii) by in situ hybridization, that OeTaq80-related DNA sequences are mainly localized in the heterochromatin at the chromosome ends; (iii) by dot-blot hybridization experiments, that these sequences are highly represented in the genome of all the olive cultivars and the majority of Olea species studied, and that their frequency may differ significantly even between olive cultivars; and (iv) by calculating the copy number of OeTaq80-related sequences per haploid (1C) genome, that the redundancy of these DNA elements may differ significantly between the genomes tested. It is suggested that the inter- and intraspecific changes in the nuclear and genomic traits observed can contribute to the understanding of the phylogenetic relationships between Olea species and in defining parameters to be exploited in varietal identification within cultivated olives.

Base Sequence↗
PubMed1993

Mapping chromosome rearrangement breakpoints to the physical map of Caenorhabditis elegans by fluorescent in situ hybridization.

A scheme for rapidly mapping chromosome rearrangements relative to the physical map of Caenorhabditis elegans is described that is based on hybridization patterns of cloned DNA on meiotic nuclei, as visualized by fluorescent in situ hybridization. From the nearly complete physical map, DNA clones, in yeast artificial chromosomes (YACs), spanning the rearrangement breakpoint were selected. The purified YAC DNAs were first amplified by degenerate oligonucleotide-primed polymerase chain reaction, then reamplified to incorporate fluorescein dUTP or rhodamine dUTP. The site of hybridization was visualized directly (without the use of antibodies) on meiotic bivalents. This allows chromosome rearrangements to be mapped readily if the duplicated, deficient or translocated regions do not pair with a normal homologous region, because the site or sites of hybridization of the probe on meiotic prophase nuclei will be spatially distinct. The pattern, or number, of hybridization signals from probes from within, or adjacent to, the rearranged region of the genome can be predicted from the genetic constitution of the strain. Characterization of the physical extent of the genetically mapped rearrangements places genetic landmarks on the physical map, and so provides linkage between the two types of map.

Animals↗