Academic literature on the topic 'Chromosome contacts'
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Journal articles on the topic "Chromosome contacts"
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Matveevsky, Sergey, Oxana Kolomiets, Aleksey Bogdanov, Elena Alpeeva, and Irina Bakloushinskaya. "Meiotic Chromosome Contacts as a Plausible Prelude for Robertsonian Translocations." Genes 11, no. 4 (April 2, 2020): 386. http://dx.doi.org/10.3390/genes11040386.
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Robertsonian translocations are common chromosomal alterations. Chromosome variability affects human health and natural evolution. Despite the significance of such mutations, no mechanisms explaining the emergence of such translocations have yet been demonstrated. Several models have explored possible changes in interphase nuclei. Evidence for non-homologous chromosomes end joining in meiosis is scarce, and is often limited to uncovering mechanisms in damaged cells only. This study presents a primarily qualitative analysis of contacts of non-homologous chromosomes by short arms, during meiotic prophase I in the mole vole, Ellobius alaicus, a species with a variable karyotype, due to Robertsonian translocations. Immunocytochemical staining of spermatocytes demonstrated the presence of four contact types for non-homologous chromosomes in meiotic prophase I: (1) proximity, (2) touching, (3) anchoring/tethering, and (4) fusion. Our results suggest distinct mechanisms for chromosomal interactions in meiosis. Thus, we propose to change the translocation mechanism model from ‘contact first’ to ‘contact first in meiosis’.
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George, Phillip, Nicholas A. Kinney, Jiangtao Liang, Alexey V. Onufriev, and Igor V. Sharakhov. "Three-dimensional Organization of Polytene Chromosomes in Somatic and Germline Tissues of Malaria Mosquitoes." Cells 9, no. 2 (February 1, 2020): 339. http://dx.doi.org/10.3390/cells9020339.
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Spatial organization of chromosome territories and interactions between interphase chromosomes themselves, as well as with the nuclear periphery, play important roles in epigenetic regulation of the genome function. However, the interplay between inter-chromosomal contacts and chromosome-nuclear envelope attachments in an organism’s development is not well-understood. To address this question, we conducted microscopic analyses of the three-dimensional chromosome organization in malaria mosquitoes. We employed multi-colored oligonucleotide painting probes, spaced 1 Mb apart along the euchromatin, to quantitatively study chromosome territories in larval salivary gland cells and adult ovarian nurse cells of Anopheles gambiae, An. coluzzii, and An. merus. We found that the X chromosome territory has a significantly smaller volume and is more compact than the autosomal arm territories. The number of inter-chromosomal, and the percentage of the chromosome–nuclear envelope, contacts were conserved among the species within the same cell type. However, the percentage of chromosome regions located at the nuclear periphery was typically higher, while the number of inter-chromosomal contacts was lower, in salivary gland cells than in ovarian nurse cells. The inverse correlation was considerably stronger for the autosomes. Consistent with previous theoretical arguments, our data indicate that, at the genome-wide level, there is an inverse relationship between chromosome-nuclear envelope attachments and chromosome–chromosome interactions, which is a key feature of the cell type-specific nuclear architecture.
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Miermans, Christiaan A., and Chase P. Broedersz. "Bacterial chromosome organization by collective dynamics of SMC condensins." Journal of The Royal Society Interface 15, no. 147 (October 2018): 20180495. http://dx.doi.org/10.1098/rsif.2018.0495.
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A prominent organizational feature of bacterial chromosomes was revealed by Hi-C experiments, indicating anomalously high contacts between the left and right chromosomal arms. These long-range contacts have been attributed to various nucleoid-associated proteins, including the ATPase Structural Maintenance of Chromosomes (SMC) condensin. Although the molecular structure of these ATPases has been mapped in detail, it still remains unclear by which physical mechanisms they collectively generate long-range chromosomal contacts. Here, we develop a computational model that captures the subtle interplay between molecular-scale activity of slip-links and large-scale chromosome organization. We first consider a scenario in which the ATPase activity of slip-links regulates their DNA-recruitment near the origin of replication, while the slip-link dynamics is assumed to be diffusive. We find that such diffusive slip-links can collectively organize the entire chromosome into a state with aligned arms, but not within physiological constraints. However, slip-links that include motor activity are far more effective at organizing the entire chromosome over all length-scales. The persistence of motor slip-links at physiological densities can generate large, nested loops and drive them into the bulk of the DNA. Finally, our model with motor slip-links can quantitatively account for the rapid arm–arm alignment of chromosomal arms observed in vivo .
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Socol, Marius, Renjie Wang, Daniel Jost, Pascal Carrivain, Cédric Vaillant, Eric Le Cam, Vincent Dahirel, et al. "Rouse model with transient intramolecular contacts on a timescale of seconds recapitulates folding and fluctuation of yeast chromosomes." Nucleic Acids Research 47, no. 12 (May 22, 2019): 6195–207. http://dx.doi.org/10.1093/nar/gkz374.
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Abstract DNA folding and dynamics along with major nuclear functions are determined by chromosome structural properties, which remain, thus far, elusive in vivo. Here, we combine polymer modeling and single particle tracking experiments to determine the physico-chemical parameters of chromatin in vitro and in living yeast. We find that the motion of reconstituted chromatin fibers can be recapitulated by the Rouse model using mechanical parameters of nucleosome arrays deduced from structural simulations. Conversely, we report that the Rouse model shows some inconsistencies to analyze the motion and structural properties inferred from yeast chromosomes determined with chromosome conformation capture techniques (specifically, Hi-C). We hence introduce the Rouse model with Transient Internal Contacts (RouseTIC), in which random association and dissociation occurs along the chromosome contour. The parametrization of this model by fitting motion and Hi-C data allows us to measure the kinetic parameters of the contact formation reaction. Chromosome contacts appear to be transient; associated to a lifetime of seconds and characterized by an attractive energy of –0.3 to –0.5 kBT. We suggest attributing this energy to the occurrence of histone tail-DNA contacts and notice that its amplitude sets chromosomes in ‘theta’ conditions, in which they are poised for compartmentalization and phase separation.
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Benedetti, Fabrizio, Julien Dorier, Yannis Burnier, and Andrzej Stasiak. "Models that include supercoiling of topological domains reproduce several known features of interphase chromosomes." Nucleic Acids Research 42, no. 5 (December 22, 2013): 2848–55. http://dx.doi.org/10.1093/nar/gkt1353.
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Abstract Understanding the structure of interphase chromosomes is essential to elucidate regulatory mechanisms of gene expression. During recent years, high-throughput DNA sequencing expanded the power of chromosome conformation capture (3C) methods that provide information about reciprocal spatial proximity of chromosomal loci. Since 2012, it is known that entire chromatin in interphase chromosomes is organized into regions with strongly increased frequency of internal contacts. These regions, with the average size of ∼1 Mb, were named topological domains. More recent studies demonstrated presence of unconstrained supercoiling in interphase chromosomes. Using Brownian dynamics simulations, we show here that by including supercoiling into models of topological domains one can reproduce and thus provide possible explanations of several experimentally observed characteristics of interphase chromosomes, such as their complex contact maps.
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Kranas, Hanna, Irina Tuszynska, and Bartek Wilczynski. "HiCEnterprise: identifying long range chromosomal contacts in Hi-C data." PeerJ 9 (April 26, 2021): e10558. http://dx.doi.org/10.7717/peerj.10558.
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Motivation Computational analysis of chromosomal contact data is currently gaining popularity with the rapid advance in experimental techniques providing access to a growing body of data. An important problem in this area is the identification of long range contacts between distinct chromatin regions. Such loops were shown to exist at different scales, either mediating relatively short range interactions between enhancers and promoters or providing interactions between much larger, distant chromosome domains. A proper statistical analysis as well as availability to a wide research community are crucial in a tool for this task. Results We present HiCEnterprise, a first freely available software tool for identification of long range chromatin contacts not only between small regions, but also between chromosomal domains. It implements four different statistical tests for identification of significant contacts for user defined regions or domains as well as necessary functions for input, output and visualization of chromosome contacts. Availability The software and the corresponding documentation are available at: github.com/regulomics/HiCEnterprise. Supplementary information Supplemental data are available in the online version of the article and at the website regulomics.mimuw.edu.pl/wp/hicenterprise.
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Orlov, Y. L., O. Thierry, A. G. Bogomolov, A. V. Tsukanov, E. V. Kulakova, E. R. Galieva, A. O. Bragin, and G. Li. "Computer methods of analysis of chromosome contacts in the cell nucleus based on sequencing technology data." Biomeditsinskaya Khimiya 63, no. 5 (2017): 418–22. http://dx.doi.org/10.18097/pbmc20176305418.
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The study spatial chromosome structure and chromosome folding in the interphase cell nucleus is an important challenge of world science. Detection of eukaryotic genome regions that physically interact with each other could be done by modern sequencing technologies. A basic method of chromosome folding by total sequencing of contacting DNA fragments is HI-C. Long-range chromosomal interactions play an important role in gene transcription and regulation. The study of chromosome interactions, 3D (three-dimensional) genome structure and its effect on gene transcription allows revealing fundamental biological processes from a viewpoint of structural regulation and are important for cancer research. The technique of chromatin immunoprecipitation and subsequent sequencing (ChIP-seq) make possible to determine binding sites of transcription factors that regulate expression of eukaryotic genes; genome transcription factors binding maps have been. The ChIA-PET technology allows exploring not only target protein binding sites, but also pairs of such sites on proximally located and interacting with each other chromosomes co-located in three-dimensional space of the cell nucleus. Here we discuss the principles of the construction of genomic maps and matrices of chromosome contacts according to ChIA-PET and Hi-C data that capture the chromosome conformation and overview existing software for 3D genome analysis including in house programs of gene location analysis in topological domains.
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Loidl, Josef. "The initiation of meiotic chromosome pairing: the cytological view." Genome 33, no. 6 (December 1, 1990): 759–78. http://dx.doi.org/10.1139/g90-115.
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Opposing views are held with respect to the time when and the mechanisms whereby homologous chromosomes find each other for meiotic synapsis. On the one hand, some evidence has been presented for somatic homologous associations or some other kind of relationship between chromosomes in somatic cells as a preliminary to meiotic pairing. On the other hand, it is argued by many that homologous contacts are first established at meiotic prophase prior to, or in the course of, synaptonemal complex formation. The present paper reviews the controversial cytological evidence, hypotheses, and ideas on how the first contact between homologous chromosomes comes about.Key words: synapsis, meiosis, presynaptic alignment, homologous recognition, synaptonemal complex, chromosome pairing.
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Stodola, Timothy J., Pengyuan Liu, Yong Liu, Andrew K. Vallejos, Aron M. Geurts, Andrew S. Greene, and Mingyu Liang. "Genome-wide map of proximity linkage to renin proximal promoter in rat." Physiological Genomics 50, no. 5 (May 1, 2018): 323–31. http://dx.doi.org/10.1152/physiolgenomics.00132.2017.
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A challenge to understanding enhancer-gene relationships is that enhancers are not always sequentially close to the gene they regulate. Physical proximity mapping through sequencing can provide an unbiased view of the chromatin close to the proximal promoter of the renin gene ( Ren). Our objective was to determine genomic regions that physically interact with the renin proximal promoter, using two different genetic backgrounds, the Dahl salt sensitive and normotensive SS-13BN, which have been shown to have different regulation of plasma renin in vivo. The chromatin conformation capture method with sequencing focused at the Ren proximal promoter in rat-derived cardiac endothelial cells was used. Cells were fixed, chromatin close to the Ren promoter was captured, and fragments were sequenced. The clustering of mapped reads produced a genome-wide map of chromatin in contact with the Ren promoter. The largest number of contacts was found on chromosome 13, the chromosome with Ren, and contacts were found on all other chromosomes except chromosome X. These contacts were significantly enriched with genes positively correlated with Ren expression and with mapped quantitative trait loci associated with blood pressure, cardiovascular, and renal phenotypes. The results were reproducible in an independent biological replicate. The findings reported here represent the first map between a critical cardiovascular gene and physical interacting loci throughout the genome and will provide the basis for several new directions of research.
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Jian, Xing, and Gary Felsenfeld. "Insulin promoter in human pancreatic β cells contacts diabetes susceptibility loci and regulates genes affecting insulin metabolism." Proceedings of the National Academy of Sciences 115, no. 20 (April 30, 2018): E4633—E4641. http://dx.doi.org/10.1073/pnas.1803146115.
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Both type 1 and type 2 diabetes involve a complex interplay between genetic, epigenetic, and environmental factors. Our laboratory has been interested in the physical interactions, in nuclei of human pancreatic β cells, between the insulin (INS) gene and other genes that are involved in insulin metabolism. We have identified, using Circularized Chromosome Conformation Capture (4C), many physical contacts in a human pancreatic β cell line between the INS promoter on chromosome 11 and sites on most other chromosomes. Many of these contacts are associated with type 1 or type 2 diabetes susceptibility loci. To determine whether physical contact is correlated with an ability of the INS locus to affect expression of these genes, we knock down INS expression by targeting the promoter; 259 genes are either up or down-regulated. Of these, 46 make physical contact with INS. We analyze a subset of the contacted genes and show that all are associated with acetylation of histone H3 lysine 27, a marker of actively expressed genes. To demonstrate the usefulness of this approach in revealing regulatory pathways, we identify from among the contacted sites the previously uncharacterized gene SSTR5-AS1 and show that it plays an important role in controlling the effect of somatostatin-28 on insulin secretion. These results are consistent with models in which clustering of genes supports transcriptional activity. This may be a particularly important mechanism in pancreatic β cells and in other cells where a small subset of genes is expressed at high levels.
Dissertations / Theses on the topic "Chromosome contacts"
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Conin, Brenna. "Genomic contacts reveal the control of sister chromosome decatenation in E. coli." Electronic Thesis or Diss., Sorbonne université, 2020. http://www.theses.fr/2020SORUS376.
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La Topo IV est responsable de la résolution des caténanes formés lors de la réplication du chromosome et joue un rôle essentiel dans la ségrégation des chromosomes. Des études antérieures ont montrées que, les altérations de Topo IV (parEts mutant), entraînent une interaction prolongée entre les chromatides sœurs, entraînant une mauvaise ségrégation et une perte de viabilité. En utilisant la capture de conformation chromosomique (Hi-C) et de microscopie à fluorescence, nous avons montré que l'altération de Topo IV affecte l'organisation de l'ensemble du chromosome. Le phénotype le plus frappant est l'émergence de deux signaux spécifiques à 1,35 Mb et 1,75 Mb. Ces loci peuvent entrer en contact avec n'importe quel locus de la partie proximale de l’origine de réplication du chromosome (cela se manifeste par des motifs caractéristiques sur les cartes de Hi-C, que nous avons appelé « ailes de papillon »). De plus, les cellules mutantes ont montré une perte de contacts dans le macrodomaine Ter, par rapport aux cellules sauvages, suggérant un changement dans l'organisation de celui-ci. Nous avons également observé une augmentation générale des contacts à courte distance le long du génome. Ces observations suggèrent qu'en l'absence de Topo IV, il y a une accumulation de precatenanes dans tout le chromosome, permettant aux loci sur différentes chromatides d'interagir (contact inter-chromosomique). Cette hypothèse est étayée par l'étude de l'interaction entre la Topo IV et la Topo III, qui a montré que la Topo III agit sur les précaténanes à une très courte distance de la fourche de réplication et ne peut pas « atteindre » les précaténanes responsables des ailes de papillon. Nous avons, également montré que la position des ailes de papillon dépend de matS et MatP. Il est intéressant de noter, que le Hi-C du double mutant matP parEts ne présente pas les signaux caractéristiques du simple mutant parEts à la frontière du ter, mais révèle plutôt que le domaine Ter lui-même est capable d’interagir avec des loci distants du chromosome. Ceci suggère que les précaténanes n'ont pas pu passer le site dif probablement à cause du complexe MatP-matS. De plus, des expériences NorFlIP précédentes ont montrés deux sites de liaison à la Topo IV à 1,2 Mb et 1,8 Mb qui ne sont pas clivés et qui s’alignent avec le centre des ailes de papillon. Nous avons ainsi émis l'hypothèse que le complexe matS-MatP et les sites Topo IV définissent un centre de décaténation. Les précaténanes non résolues seraient « attirées » vers ce centre, pour être décaténées avant la division cellulaire. Dans cette hypothèse, la liaison entre le Ter et le divisome (« Ter linkage ») joue un rôle essentiel dans le centre de décaténation car elle empêche les précaténanes de passer par dif. L'absence de Topo IV fonctionnelle va donc perturber le centre de décaténation, se traduisant par une accumulation de précaténanes en bordure du signal du centre estropié et des ailes de papillon sur une matrice Hi-C. Au regard de cette hypothèse, nous avons étudié le rôle de MukB capable de condenser l'ADN, éventuellement par extrusion en boucle, et montré que MukB définit la longueur et la densité des ailes du papillonTopoisomerase IV, is responsible for the untangling of catenanes that are formed during the replication of the chromosome and has been shown to play an essential role in nucleoid segregation. Previous studies have shown that alterations in Topo IV result in a prolonged interaction between sister chromosomes leading to poor chromosome segregation and a loss in cell viability. Using chromosome conformation capture (Hi-C) and fluorescence microscopy, we have shown that the alteration of Topo IV affects the organisation of the entire chromosome. The most striking phenotype is the emergence of two distinct signals at 1.35Mb and 1.75Mb where loci in these regions are able to contact any loci of the origin-proximal part of the chromosome (butterfly wings). Furthermore, when compared to WT cells, the mutant cells showed a loss of contacts within the terminus domain, suggesting a change in the organisation of the ter domain. We also observed a general increase of short-range contacts along the diagonal. This phenotype was only observed in E. coli cells with a circular chromosome that was undergoing replication. Those observations suggest that in the absence of Topo IV, there is an accumulation of precatenanes throughout the chromosome, allowing loci on different siter chromosomes to interact (inter-chromosomal contacts). This hypothesis was further supported when we studied the interplay between Topo IV and Topo III, which showed that Topo III acts on precatenanes at a very short distances from the replication fork and cannot “reach” precatenanes responsible for the butterfly wing signals. We further showed that the butterfly wing positions are dependent on both matS and MatP. Interestingly, Hi-C of the matP parEts double mutant does not display the characteristic signals of the single parEts mutant at the border of the ter, but instead reveals that the ter domain itself is able to contact distant loci of the chromosome. This suggests that the precatenanes were unable to go passed the dif site probably because of the MatP-matS complex. In addition, previous NorFlIP experiments have showen that Topo IV is able to bind but not cleave at two sites positioned at 1.2Mb and 1.8Mb, which align with the centre of the butterfly wings. We thereby hypothesised that the matS-MatP complex and these Topo IV sites define a decatenation hub. Unresolved precatenanes would be “pulled” toward this hub, to be decatenated prior to cell division. In this hypothesis, the Ter linkage plays an essential role in the decatenation hub as it prevents precatenanes from passing through dif. The absence of a functional Topo IV will therefore disturb the decatenation hub, resulting in accumulation of precatenanes at the border of the crippled hub and this is turn would be represented as the butterfly wing signals seen on a Hi-C matrix. In regard to this hypothesis, we investigated the role of MukB that is able to condense the DNA, possibly by loop extrusion, and show that MukB defines the length and density of the butterfly wings
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Girard, Fabien. "Tethering of molecular parasites on inactive chromatin in eukaryote nucleus." Electronic Thesis or Diss., Sorbonne université, 2023. http://www.theses.fr/2023SORUS661.
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Les plasmides naturels sont courants chez les procaryotes, mais peu ont été documentés chez les eucaryotes. Le plasmide naturel 2µ présent dans la levure bourgeonnante Saccharomyces cerevisiae est l'un des mieux caractérisés. Cet élément génétique très stable coexiste avec son hôte depuis des millions d'années, ségrégeant efficacement à chaque division cellulaire par un mécanisme qui reste mal compris. En utilisant la ligature de proximité (Hi-C, Micro-C) pour cartographier les contacts entre le plasmide 2µ et les chromosomes de levure dans des dizaines de conditions biologiques différentes, nous avons constaté que le plasmide 2µ se fixe préférentiellement sur des régions à faible activité transcriptionnelle, correspondant souvent à de longs gènes inactifs. Les acteurs communs de la structure des chromosomes, tels que les membres des complexes de maintenance structurale des chromosomes (SMC), ne sont pas impliqués dans ces contacts qui dépendent plutôt d'un signal nucléosomique associé à une déplétion de l'ARN Pol II. Ces contacts sont stables tout au long du cycle cellulaire et peuvent être établis en quelques minutes. Cette stratégie peut aussi être trouvée dans d'autres types de molécules d'ADN et d'autres espèces que S. cerevisiae, comme le suggère le schéma de liaison du plasmide naturel le long des régions silencieuses des chromosomes de Dictyostelium discoideumNatural plasmids are common in prokaryotes but few have been documented in eukaryotes. The natural 2µ plasmid present in budding yeast Saccharomyces cerevisiae is one of the most well characterized. This highly stable genetic element coexists with its host for millions of years, efficiently segregating at each cell division through a mechanism that remains poorly understood. Using proximity ligation (Hi-C, MicroC) to map the contacts between the 2µ and yeast chromosomes under dozens of different biological conditions, we found that the plasmid tether preferentially on regions with low transcriptional activity, often corresponding to long inactive genes, throughout the cell cycle. Common players in chromosome structure such as members of the structural maintenance of chromosome complexes (SMC) are not involved in these contacts, and depend instead on a nucleosomal signal associated with a depletion of RNA Pol II. These contacts are highly stable, and can be established within minutes. Our data show that the plasmid segregates by binding to transcriptionally silent regions of the host chromosomes. This strategy may concern other types of DNA molecules and species beyond S. cerevisiae, as suggested by the binding pattern of the natural Ddp5 plasmid along Dictyostelium discoideum chromosomes’ silent regions
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Carron, Léopold. "Analyse à haute résolution de la structure spatiale des chromosomes eucaryotes Boost-HiC : Computational enhancement of long-range contacts in chromosomal contact maps Genome supranucleosomal organization and genetic susceptibility to disease." Thesis, Sorbonne université, 2019. http://www.theses.fr/2019SORUS593.
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L’information génétique est portée par la molécule d’ADN, un polymère de nucléotides de très grande taille. Afin de mieux comprendre les mécanismes impactant le repliement de l’ADN, on peut exploiter une technique de génomique qui permet de quantifier les contacts entre régions distales du génome. Cette technique expérimentale appelée ’capture de conformation de chromosome’ (Hi-C) donne des informations quantitatives sur l’architecture et le repliement tridimensionnel des chromosomes dans le noyau. Largement utilisée chez l’Homme, la souris et la drosophile, cette technique a grandement évolué durant ces dernières années, produisant ainsi des données de qualité variable. Jusque-là étudiées à des résolutions assez grossières, notre objectif est d’étudier les données Hi-C déjà publiées à des résolutions plus fines. Pour cela, j’ai développé un outil bioinformatique, Boost-HiC, pour améliorer l’analyse des contacts chromosomiques. Fort de cette expertise, je proposerai alors une analyse comparative des structures spatiales des génomes eucaryotes, permettant de clarifier comment extraire les compartiments génomiques de manière optimale. Cette expertise sera utilisée également pour décrire le lien entre les bordures des domaines topologiques de la chromatine et la position dans le génome humain des mutations ponctuelles prédisposant au cancerGenetic information is encoded in DNA, a huge-size nucleotidic polymer. In order to understand DNA folding mechanisms, an experimental technique is today available that quantifies distal genomic contacts. This high-throughput chromosome conformation capture technique, called Hi-C, reveals 3D chromosome folding in the nucleus. In the recent years, the Hi-C experimental protocol received many improvements through numerous studies for Human, mouse and drosophila genomes. Because most of these studies are performed at poor resolution, I propose bioinformatic methods to analyze these datasets at fine resolution. In order to do this, I present Boost-HiC, a tool that enhanced long-range contacts in Hi-C data. I will then used our extended knowledge to compare 3D folding in different species. This result provides the basis to determine the best method for obtaining genomic compartements from a chromosomal contact map. Finally, I present some other applications of our methodology to study the link between the borders of topologically associating domains and the genomic location of single-nucleotide mutations associated to cancer
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Riley, Anthony David. "Probing chromosome structure using multidimensional scaling of DNA contact matrices." Thesis, University of Leeds, 2014. http://etheses.whiterose.ac.uk/7262/.
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Chromosome conformation capture technology has provided a route to studying genome structure through DNA-DNA contact-counts. An iteration of chromosome conformation capture technology is Hi-C, which provides genome wide two dimensional contact-count data. The contact-count data from Hi-C can be viewed as a proxy for distance and using some transform function can be transformed into estimated distances. These estimated distances can be fitted into Euclidean space using the statistical tools of multidimensional scaling to give estimated chromosome or genome configurations. The first part of this thesis takes the Hi-C contact-count data for Chromosome 14, transforms it into estimated distances which are fitted into Euclidean space to give an estimated chromosome configuration. Steps are also taken to pre-process the genome contact-count matrix to refine the information held within it. The pre-processed genome contact-count matrix is transformed into estimated distances, which are fitted into Euclidean space to give an estimated genome configuration. The estimated chromosome and genome configurations are investigated, to find if known features of these structures are captured through fitting the Hi-C data. The second part of this thesis simulates contact-count data from simple configurations. Using the inverse of the transform functions the distances between points in a configuration can be transformed into mean contact-counts. The mean contact-counts are perturbed using a suitable distribution function to provide perturbed contact-counts, which are transformed into perturbed distances. The perturbed distances can be fitted into Euclidean space to give a fitted configurations. The properties of the fitted configurations are investigated and compared with the original configurations, and the properties of the perturbed distances are also investigated. Then steps are taken to improve the fitted configurations using information from the properties of the perturbed distances, with the successful techniques applied to estimating the chromosome configuration.
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Heurteau, Alexandre. "Etude bioinformatique intégrative : déterminants et dynamique des interactions chromosomiques à longue distance." Electronic Thesis or Diss., Toulouse 3, 2019. http://www.theses.fr/2019TOU30343.
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Les protéines se liants aux insulateurs (IBPs) seraient impliquées dans la structuration tri-dimensionnelle des génomes en domaines topologiques (ou " TADs). Les TADs contribueraient notamment à séparer les compartiments inactifs/hétérochromatine et actifs/euchromatine. Les IBPs sont également capables de bloquer les contacts spécifiques entre les éléments activateurs ou "enhancers" d'un TAD et les promoteurs de gènes cibles présents dans un autre TAD. Ainsi, les insulateurs influenceraient l'expression des gènes selon plusieurs modes de régulations qui reste à être caractérisés à l'échelle du génome. Les résultats obtenus dans la première partie de ma thèse montrent comment les IBPs influenceraient l'expression des gènes selon un nouveau mécanisme de régulation, comme montré à l'échelle du génome de la Drosophile. Nos analyses bioinformatiques montrent que les IBPs régulent l'étalement de l'hétérochromatine répressive (H3K27me3) à la fois en cis et en trans. Les régulations en trans impliquent des boucles de chromatine entre insulateurs positionnés à la frontière de l'hétérochromatine et des insulateurs distants positionnés aux abords de gènes euchromatiniens. Ces étalements en trans conduisent à la formation de "micro-domaines" d'hétérochromatine réprimant ainsi les gènes distants. En particulier, un mutant d'insulateur qui empêche la formation de boucle diminue significativement l'établissement des micro-domaines. De plus, ces micro-domaines se formeraient au cours du développement suggérant un nouveau mécanisme insulateur-dépendant de régulation des gènes. De plus, nous un nouveau rôle de la Cohésine, un régulateur clé des boucles 3D chez l'homme, dans la régulation des ARN non codants (ncRNAs), incluant les "PROMoters uPstream Transcripts" (PROMPTs) et les enhancers RNAs (eRNAs). L'hélicase MTR4 est essentielle au contrôle de la stabilité des ARNs codants et non codants par son rôle dans les complexes nuclear-exosome targeting (NEXT) et pA-tail exosome targeting (PAXT). De manière intéressante, la déplétion de MTR4 et des sous-unités ZFC3H1 et ZCCHC8 (ou Z1 et Z8), a conduit à l'apparition de ncRNAs à l'échelle du génome. Curieusement, la cartographie des sites de liaison de MTR4 a mis en évidence que cette hélicase se lie sur des sites distants des PROMPTs. Plutôt que d'agir en cis, nos données suggèrent que la régulation des PROMPTs pourrait impliquer des contacts spécifiques à longue distance entre ces sites distants de liaison MTR4 et les promoteurs liés par Z1/Z8. Ainsi, l'intégration des données Hi-C et la détection des PROMPTS en conditions de déplétion de MTR4, Z1 ou Z8 ont souligné le rôle possible des interactions à longue distance dans la régulation des PROMPTs, depuis les sites distants MTR4. Ces travaux pourraient établir une nouvelle relation entre la structure 3D des génomes et la régulation des ARNs non codantsInsulator Binding Proteins (IBPs) could be involved in the three-dimensional folding of genomes into topological domains (or "TADs"). In particular, TADs would help to separate the inactive/heterochromatin and active/euchromatin compartments. IBPs are also able to block specific contacts between the activator or enhancer elements of one TAD and target gene promoters present in another TAD. Thus, insulators may influence gene expression according to several regulatory modes that have yet to be characterized at genome level. The results obtained in the first part of my thesis show how IBPs influence gene expression according to a new regulatory mechanism, as shown at the scale of the Drosophila genome. Our bioinformatics analyses show that IBPs regulate the spread of repressive heterochromatin (H3K27me3) both in cis and trans. Trans regulations involve chromatin loops between insulators positioned at the heterochromatin boundary and distant insulators positioned at the edges of euchromatic genes. Trans spreading leads to the formation of "micro-domains" of heterochromatin, thereby repressing distant genes. In particular, an insulator mutant that prevents loop formation significantly reduces the establishment of micro-domains. In addition, these micro-domains would be formed during development suggesting a new insulator-dependent mechanism for gene regulation. Furthermore, we could uncover a novel function of cohesion, a key regulator of 3D loops in humans, in regulating non-coding RNAs (ncRNAs), including "PROMoters uPstream Transcripts" (PROMPTs) and enhancers RNAs (eRNAs). The MTR4 helicase is essential to the control of coding and noncoding RNA stability by the human nuclear-exosome targeting (NEXT) complex and pA-tail exosome targeting (PAXT) complex. Remarkably, ncRNAs could be detected upon depletion of the Mtr4 helicase of the human NEXT complex. Moreover, depletion of additional NEXT subunits, ZFC3H1 and ZCCHC8 (or Z1 and Z8), also led to uncover ncRNAs often produced from the same loci as upon MTR4 depletion. Curiously however, mapping of Mtr4 binding sites highlighted that Mtr4 binds to sites that are distant from PROMPTs. Rather than acting in cis, our data suggest that regulation of PROMPTs could involve specific long-distance contacts between these distant MTR4 binding sites and promoters bound by Z1/Z8. As such, integration of Hi-C data together with the detection of PROMPTS upon MTR4-, Z1- or Z8- depletions highlight possible role of long-range interactions in regulating PROMPTs, from distant MTR4-bound sites. This work may establish a new relationship between the 3D structure of genomes and the regulation of ncRNAs
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Le, Treut Guillaume. "Models of chromosome architecture and connection with the regulation of genetic expression." Thesis, Université Paris-Saclay (ComUE), 2016. http://www.theses.fr/2016SACLS411/document.
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Plusieurs indices suggèrent que le repliement du chromosome et la régulation de l’expression génétique sont étroitement liés. Par exemple, la co-expression d’un grand nombre de gènes est favorisée par leur rapprochement dans l’espace cellulaire. En outre, le repliement du chromosome permet de faire émerger des structures fonctionnelles. Celles-ci peuvent être des amas condensés et fibrillaires, interdisant l’accès à l’ADN, ou au contraire des configurations plus ouvertes de l’ADN avec quelques amas globulaires, comme c’est le cas avec les usines de transcription. Bien que dissemblables au premier abord, de telles structures sont rendues possibles par l’existence de protéines bivalentes, capable d’apparier des régions parfois très éloignées sur la séquence d’ADN. Le système physique ainsi constitué du chromosome et de protéines bivalentes peut être très complexe. C’est pourquoi les mécanismes régissant le repliement du chromosome sont restés majoritairement incompris.Nous avons étudié des modèles d’architecture du chromosome en utilisant le formalisme de la physique statistique. Notre point de départ est la représentation du chromosome sous la forme d’un polymère rigide, pouvant interagir avec une solution de protéines liantes. Les structures résultant de ces interactions ont été caractérisées à l’équilibre thermodynamique. De plus, nous avons utilisé des simulations de dynamique Brownienne en complément des méthodes théoriques, car elles permettent de prendre en considération une plus grande complexité dans les phénomènes biologiques étudiés.Les principaux aboutissements de cette thèse ont été : (i) de fournir un modèle pour l’existence des usines de transcriptions caractérisées in vivo à l’aide de microscopie par fluorescence ; (ii) de proposer une explication physique pour une conjecture portant sur un mécanisme de régulation de la transcription impliquant la formation de boucles d’ADN en tête d’épingle sous l’effet de la protéine H-NS, qui a été émise suite à l’observation de ces boucles au microscope à force atomique ; (iii) de proposer un modèle du chromosome qui reproduise les contacts mesurés à l’aide des techniques Hi-C. Les conséquences de ces mécanismes sur la régulation de la transcription ont été systématiquement discutéesIncreasing evidences suggest that chromosome folding and genetic expression are intimately connected. For example, the co-expression of a large number of genes can benefit from their spatial co-localization in the cellular space. Furthermore, functional structures can result from the particular folding of the chromosome. These can be rather compact bundle-like aggregates that prevent the access to DNA, or in contrast, open coil configurations with several (presumably) globular clusters like transcription factories. Such phenomena have in common to result from the binding of divalent proteins that can bridge regions sometimes far away on the DNA sequence. The physical system consisting of the chromosome interacting with divalent proteins can be very complex. As such, most of the mechanisms responsible for chromosome folding and for the formation of functional structures have remained elusive.Using methods from statistical physics, we investigated models of chromosome architecture. A common denominator of our approach has been to represent the chromosome as a polymer with bending rigidity and consider its interaction with a solution of DNA-binding proteins. Structures entailed by the binding of such proteins were then characterized at the thermodynamical equilibrium. Furthermore, we complemented theoretical results with Brownian dynamics simulations, allowing to reproduce more of the biological complexity.The main contributions of this thesis have been: (i) to provide a model for the existence of transcrip- tion factories characterized in vivo with fluorescence microscopy; (ii) to propose a physical basis for a conjectured regulatory mechanism of the transcription involving the formation of DNA hairpin loops by the H-NS protein as characterized with atomic-force microscopy experiments; (iii) to propose a physical model of the chromosome that reproduces contacts measured in chromosome conformation capture (CCC) experiments. Consequences on the regulation of transcription are discussed in each of these studies
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Moran, Peter. "A behavioural and genomic approach to studying the evolution of reproductive isolation : a contact zone between closely related field crickets in the genus Teleogryllus." Thesis, University of St Andrews, 2017. http://hdl.handle.net/10023/10260.
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What processes contribute to the evolution of reproductive isolation and the coexistence of interfertile species in the same habitat? This thesis investigates the relative roles of species interactions and intraspecific processes in contributing to reproductive isolation. I combine behavioural and genomic approaches to test hypotheses about what mechanisms maintain the general species boundary between two closely related field cricket species: Teleogryllus oceanicus and T. commodus. These species are a classic study system for sexual communication and readily hybridize in the laboratory, however little is known about species interactions in sympatric populations. I examine patterns of geographic variation in two key sexual traits: calling song and cuticular hydrocarbons (CHCs), and the geographic distribution of genetic variation across a broad sample of allopatric and sympatric populations. I test whether X chromosomes play a pronounced role in population divergence and reproductive isolation. Using close range mating trials and hybridization experiments I identify numerous pre-mating and post-mating barriers between the species. The results indicate that the species are currently reproductively isolated and the pattern of population differentiation does not strongly support contemporary species interactions contributing to phenotypic diversity. Numerous barriers exist between the species, in particular hybrid females are sterile in both cross directions, while hybrid males are relatively fertile. This provides a rare exception to Haldane's rule which is central to many genetic theories of speciation. Established theory predicts that X chromosomes should play a pronounced role in the evolution of both pre- and postzygotic barriers. Contrary to this, I found no evidence that X chromosomes contribute to hybrid female sterility. Moreover, X-linked loci exhibited an unexpected pattern of reduced population differentiation within species, but increased species divergence compared to autosomal loci, which may indicate selective sweeps or sex-biased processes. Taken together, the results suggest that the causes and consequences of X chromosome evolution, in particular among XO taxa, may contradict some of the established theories.
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Zaborowski, Rafał. "Computational methods for differential analysis of chromatin contact matrices." Doctoral thesis, 2021. https://depotuw.ceon.pl/handle/item/3868.
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Understanding the relationships between chromatin structure and gene regulation is a fundamental problem in genetics. However, for a long time there has been little progress in the field of genome architecture except for studying low scale chromatin organization. This situation changed during last two decades due to the advancement in the development of NGS technology, which gave rise to Chromosome Conformation
Capture (3C) methods. The availability of 3C techniques enabled genome organization studies on an unprecedented scale. In particular, the 3C-derived Hi-C protocol allowed researchers to interrogate millions of chromatin interactions between pairs of regions genome-wide at a very high resolution. One of the main applications of Hi-C is the differential analysis, which aim to identify the structural differences of chromatin influencing regulatory processes across various cell types, treatments or species.
In this thesis we focus on the issue of comparing Hi-C contact matrices. First, we study the problem of assessing the similarity between chromosome segmentations arising from identification of Topologically Associating Domains (TAD) - an inherent feature of mammalian Hi-C maps, which were shown to shape regulatory landscape of the genome. We present a novel distance measure called BP-score, tailored for
comparison of TAD partitionings and prove that our measure satisfy metric properties. Evaluation of the BP-score on real and simulated datasets demonstrates that it performs competitive against existing approaches. Additionally, we introduce local measures of domain rearrangement and show their correlation with functional measurements.
Second, we develop a normalization-free method for discovery of Hi-C differential interactions called DiADeM. Our method introduces an intuitive definition of differential interaction, which takes into account the cross-dataset contact profile similarity. Finally, we assess DiADeMs ability to detect differential interactions using simulated contact maps and show it performs well against other available methods for Hi-C differential analysis. In summary, the tools developed by us may help researches in discovering unknown structural alterations driving regulatory mechanisms.Problem zrozumienia relacji pomiędzy strukturą chromatyny, a regulacją genów ma kluczowe znaczenie w genetyce. Niestety przez wiele lat możliwe były wyłącznie badania architektury genomu w niskiej rozdzielczości lub małej skali. Sytuacja zmieniła się w ciągu ostatnich dwóch dekad głównie ze względu na postęp w rozwoju technologii NGS, która dała początek metodom 3C (ang. Chromosome Conformation Capture). Dostępność technik 3C umożliwiła badania organizacji genomu na niespotykaną dotąd skalę. W szczególności wysoko-rozdzielczy wariant metody 3C - protokół Hi-C pozwala uzyskać dane dotyczące milionów interakcji pomiędzy parami regionów chromatyny w całym genomie. Jednym z głównych zastosowań protokołu Hi-C jest analiza różnicowa, która ma na celu zidentyfikowanie różnic w strukturze chromatyny wpływających na procesy regulacji genów w różnych typach komórek, warunkach eksperymentalnych lub gatunkach. W tej pracy koncentrujemy się na problemie porównywania macierzy kontaktów Hi-C. Po pierwsze, badamy problem oceny podobieństwa między segmentacjami chromosomów wynikającymi z identyfikacji domen topologicznych (TAD) - nieodłącznej cechy map Hi-C organizmów ssaków, które, jak wykazano, kształtują krajobraz regulacyjny genomu. Prezentujemy nową miarę odległości o nazwie BP-score, dostosowaną do porównania segmentacji TAD oraz dowodzimy, że nasza miara jest metryką. Przykładowe analizy porównawcze przeprowadzone na danych symulowanych i rzeczywistych pokazują, że odległość BP jest konkurencyjna w stosunku do innych metryk wykorzystywanych dotychczas podczas badania podobieństwa segmentacji. Dodatkowo wprowadzamy lokalne miary rearanżacji domen topologicznych i pokazujemy, że pomiary rearanżacji uzyskane przy użyciu wprowadzonych przez nas miar korelują z pomiarami ekspresji genów lub metylacji. Po drugie, opracowujemy metodę do wykrywania różnicowych oddziaływań HiC o nazwie DiADeM działającą na danych nieznormalizowanych. Nasza metoda wprowadza intuicyjną definicję interakcji różnicowych, która uwzględnia podobieństwo profili kontaktów pomiędzy zestawami danych. Na koniec oceniamy zdolność naszej metody do wykrywania interakcji różnicowych przy użyciu symulowanych map kontaktów i pokazujemy, że osiąga konkurencyjne wyniki w porównaniu z innymi dostępnymi metodami służącymi do analizy różnicowej Hi-C. Podsumowując, opracowane przez nas narzędzia mogą pomóc badaczom w odkrywaniu nieznanych zmian strukturalnych wpływających na mechanizmy regulacji genów.
Books on the topic "Chromosome contacts"
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Tral’, Tat’yana, Gulrukhsor Tolibova, Igor Kogan, and Anna Olina. Embryo losses. Atlas. ru: Publishing Center RIOR, 2023. http://dx.doi.org/10.29039/978-5-907218-78-9.
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Histologic examination of abortive material is the basic approach to identify the etiology of miscarriage. Morphological diagnostics in case of embryo loss makes it possible to draw up the plan to fully prepare the woman for future pregnancy, whether spontaneous or after fertility treatment, increasing the chance of a favorable outcome. This educational book contains the data from various studies of the endometrium and abortive material undertaken at the Ott Research Institute of Obstetrics, Gynecology and Reproductology. Histology illustrations are supplemented with images of immunohistochemical studies and confocal laser scanning microscopy photos, as well as detailed text descriptions. Images can be viewed in the atlas, with QR codes linking to high-resolution electronic photos. This edition highlights the features of endometrial structural changes related to different modes of conception, the details of assessing abortive material, trophoblast chromosomal abnormalities, anembryony, hydatidiform mole, choriocarcinoma, as well as examination of embryo losses of various origins. The atlas is intended for pathologists, obstetrician-gynecologists and heads of women’s health clinics, perinatal centers, gynecological departments of general hospitals, fertility specialists, clinical laboratory diagnostics specialists, fellows and heads of departments of obstetrics and gynecology, pathological anatomy, students of all forms of continuous medical education, graduate students and clinical residents.
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McKinlay Gardner, R. J., and David J. Amor. Centromere Fissions, Complementary Isochromosomes, Telomeric Fusions, Balancing Supernumerary Chromosomes, Neocentromeres, Jumping Translocations, and Chromothripsis. Edited by R. J. McKinlay Gardner and David J. Amor. Oxford University Press, 2018. http://dx.doi.org/10.1093/med/9780199329007.003.0012.
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This chapter reviews a number of very rare chromosomal rearrangements: centromere fissions, complementary isochromosomes, telomeric fusions, balancing supernumerary chromosomes, neocentromeres, jumping translocations, and chromothripsis. Centromere fission results when a metacentric or submetacentric chromosome splits at the centromere, giving rise to two stable telocentric products. The Robertsonian fission reverses the fusion that had originally generated it. Telomeric fusion leads to a 45-chromosome count. With the balanced complementary isochromosome carrier, two stable exactly metacentric products are generated. A balancing small supernumerary marker chromosome contains material deleted from the normal homolog. A supernumerary chromosome lacking a normal centromere can become stable and functional due to the generation of a neocentromere. In jumping translocations, a segment can move from one chromosome to two or more recipient chromosomes. Chromothripsis takes complex rearrangement to a yet more complex level. The reproductive risks associated with each are noted.
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Chromosome-2023: Proceeding of the International Conference. September 5-10, 2023. Novosibirsk State University, 2023. http://dx.doi.org/10.25205/978-5-4437-1514-8.
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The Abstract book contains materials presented at the International Conference "Chromosome-2023". Main areas discussed at the conference are devoted to the organization and evolution of chromosomes and genome, heterochromatin, genetic organization of interphase chromosomes, structure of nucleus and other topics. These materials may be interested for the scientist working in the field of genetics and molecular biology.
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Capone, George T. Down Syndrome. Oxford University Press, 2017. http://dx.doi.org/10.1093/med/9780199937837.003.0056.
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People with Down syndrome (trisomy 21) are distinguished by having an extra copy of chromosome 21. Chromosome 21 contains an estimated 562 genes, including 161 known to code for functional proteins, and at least 396 considered novel. Gene dosage imbalance is the primary mechanism, which results in the molecular, cellular, histological, and anatomical features characteristic of the condition. Throughout brain development, major neurobiological events go awry, resulting in a differently organized brain and characteristic developmental delays noted during infancy and the preschool years. The consequences of gene dosage imbalance continue to have repercussions on neurobiological function throughout childhood and adult life.
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Peñagarikano, Olga, and Daniel H. Geschwind. CNTNAP2 and Autism Spectrum Disorders. Oxford University Press, 2013. http://dx.doi.org/10.1093/med/9780199744312.003.0016.
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Although autism was described in the early 1940s as a disorder of affective contact (Kanner, 1943), it was not classified as a neurodevelopmental disorder with a biological basis until the early 1980s, when studies reported its high heritability (Folstein & Rutter, 1977; Ritvo et al., 1985) and co-occurrence with chromosomal abnormalities (Gillberg & Wahlstrom, 1985; Wahlström et al., 1986). Today, autism is considered a heterogeneous neurodevelopmental syndrome and therefore termed autism spectrum disorder (ASD), characterized by variable deficits in social behavior and language, restrictive interests, and repetitive behaviors. Autism spectrum disorder has an estimated prevalence of 1:150–1:200 (Centers for Disease Control and Prevention, 2007), being one of the most common childhood disorders. In addition to the core domains necessary for diagnosis, a number of other behavioral abnormalities are frequently associated with ASD, including epilepsy, sensory abnormalities, hyperactivity, motor abnormalities, sleep disturbances, and gastrointestinal symptoms (Geschwind, 2009).
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Pediatric ICD-10-CM. American Academy of Pediatrics, 2015. http://dx.doi.org/10.1542/9781581109016.
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In this first edition, Pediatric ICD-10-CM: A Manual for Provider-Based Coding strives to bring to the pediatric provider, coder, and biller the most accurate and easy-to use manual on ICD-10-CM yet. Composed entirely with a pediatrics focus, this manual exclusively features codes and guidelines for physician- and provider-based coding, all in a simplified yet familiar format. The full draft of the ICD-10-CM code set comes in at well over 1,000 pages. This book condenses that large and potentially cumbersome book into 400 pages of the most relevant,pediatrics-related codes and guidelines. It also fully integrates into the tabular list specific chapter and code guidelines. Guideline are now included directly at the chapter and code level, ensuring that coders will always use the right codes in right circumstances Features include Integrated codes and guidelines Simplified yet familiar layout Tabular and indexed navigation Pediatric-focused and provider-based guidance And more... Contents Include: ICD-10-CM Official Guidelines for Coding and Reporting: FY 2015 Certain Infectious and Parasitic Diseases (A00-B99) Neoplasms (C00-D49) Diseases of the Blood and Blood-Forming Organs and Certain Disorders Involving the Immune Mechanism (D50-D89) Endocrine, Nutritional and Metabolic Diseases (E00-E89) Mental, Behavioral and Neurodevelopmental Disorders (F01-F99) Diseases of the Nervous System (G00-G99) Diseases of the Eye and Adnexa (H00-H59) Diseases of the Ear and Mastoid Process (H60-H95) Diseases of the Circulatory System (I00-I99) Diseases of the Respiratory System (J00-J99) Diseases of the Digestive System (K00-K95) Diseases of the Skin and Subcutaneous Tissue (L00-L99) Diseases of the Musculoskeletal System and Connective Tissue (M00-M99) Diseases of the Genitourinary System (N00-N99) Pregnancy, Childbirth, Certain Conditions Originating in the Perinatal Period (P00-P99) Congenital Malformations, Deformations and Chromosomal Abnormalities (Q00-Q99) Symptoms, Signs, and Abnormal Clinical and Laboratory Findings (R00-R99) Injury, Poisoning and Consequences of Certain Other External Causes (S00-T88) External Causes of Morbidity (V00-Y99) Factors Influencing Health Status and Contact With Health Services (Z00-Z99)
Book chapters on the topic "Chromosome contacts"
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Paro, Renato, Ueli Grossniklaus, Raffaella Santoro, and Anton Wutz. "Biology of Chromatin." In Introduction to Epigenetics, 1–28. Cham: Springer International Publishing, 2021. http://dx.doi.org/10.1007/978-3-030-68670-3_1.
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AbstractThis chapter provides an introduction to chromatin. We will examine the organization of the genome into a nucleosomal structure. DNA is wrapped around a globular complex of 8 core histone proteins, two of each histone H2A, H2B, H3, and H4. This nucleosomal arrangement is the context in which information can be established along the sequence of the DNA for regulating different aspects of the chromosome, including transcription, DNA replication and repair processes, recombination, kinetochore function, and telomere function. Posttranslational modifications of histone proteins and modifications of DNA bases underlie chromatin-based epigenetic regulation. Enzymes that catalyze histone modifications are considered writers. Conceptually, erasers remove these modifications, and readers are proteins binding these modifications and can target specific functions. On a larger scale, the 3-dimensional (3D) organization of chromatin in the nucleus also contributes to gene regulation. Whereas chromosomes are condensed during mitosis and segregated during cell division, they occupy discrete volumes called chromosome territories during interphase. Looping or folding of DNA can bring regulatory elements including enhancers close to gene promoters. Recent techniques facilitate understanding of 3D contacts at high resolution. Lastly, chromatin is dynamic and changes in histone occupancy, histone modifications, and accessibility of DNA contribute to epigenetic regulation.
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McCauley, Micah J., Joha Joshi, Nicole Becker, Qi Hu, Maria Victoria Botuyan, Ioulia Rouzina, Georges Mer, L. James Maher, and Mark C. Williams. "Quantifying ATP-Independent Nucleosome Chaperone Activity with Single-Molecule Methods." In Single Molecule Analysis, 29–55. New York, NY: Springer US, 2023. http://dx.doi.org/10.1007/978-1-0716-3377-9_2.
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AbstractThe dynamics of histone-DNA interactions govern chromosome organization and regulates the processes of transcription, replication, and repair. Accurate measurements of the energies and the kinetics of DNA binding to component histones of the nucleosome under a variety of conditions are essential to understand these processes at the molecular level. To accomplish this, we employ three specific single-molecule techniques: force disruption (FD) with optical tweezers, confocal imaging (CI) in a combined fluorescence plus optical trap, and survival probability (SP) measurements of disrupted and reformed nucleosomes. Short arrays of positioned nucleosomes serve as a template for study, facilitating rapid quantification of kinetic parameters. These arrays are then exposed to FACT (FAcilitates Chromatin Transcription), a non-ATP-driven heterodimeric nuclear chaperone known to both disrupt and tether histones during transcription. FACT binding drives off the outer wrap of DNA and destabilizes the histone-DNA interactions of the inner wrap as well. This reorganization is driven by two key domains with distinct function. FD experiments show the SPT16 MD domain stabilizes DNA-histone contacts, while the HMGB box of SSRP1 binds DNA, destabilizing the nucleosome. Surprisingly, CI experiments do not show tethering of disrupted histones, but increased rates of histone release from the DNA. SI experiments resolve this, showing that the two active domains of FACT combine to chaperone nucleosome reassembly after the timely release of force. These combinations of single-molecule approaches show FACT is a true nucleosome catalyst, lowering the barrier to both disruption and reformation.
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Matthey-Doret, Cyril, Lyam Baudry, Shogofa Mortaza, Pierrick Moreau, Romain Koszul, and Axel Cournac. "Normalization of Chromosome Contact Maps: Matrix Balancing and Visualization." In Methods in Molecular Biology, 1–15. New York, NY: Springer US, 2021. http://dx.doi.org/10.1007/978-1-0716-1390-0_1.
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Yu, Zulin, and Tamara A. Potapova. "Superresolution for Visualization of Physical Contacts Between Chromosomes at Nanoscale Resolution." In Methods in Molecular Biology, 359–75. New York, NY: Springer US, 2022. http://dx.doi.org/10.1007/978-1-0716-2140-0_20.
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Ea, Vuthy, Franck Court, and Thierry Forné. "Quantitative Analysis of Intra-chromosomal Contacts: The 3C-qPCR Method." In Methods in Molecular Biology, 75–88. New York, NY: Springer New York, 2015. http://dx.doi.org/10.1007/7651_2015_269.
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Shinohara, K., H. Nakano, M. Watanabe, Y. Kinjo, S. Kikuchi, Y. Kagoshima, K. Kobayashi, and H. Maezawa. "X-Ray Contact Microscopy of Human Chromosomes and Human Fibroblasts." In Springer Series in Optical Sciences, 429–32. Berlin, Heidelberg: Springer Berlin Heidelberg, 1988. http://dx.doi.org/10.1007/978-3-540-39246-0_77.
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Marbouty, Martial, and Romain Koszul. "Generation and Analysis of Chromosomal Contact Maps of Bacteria." In The Bacterial Nucleoid, 75–84. New York, NY: Springer New York, 2017. http://dx.doi.org/10.1007/978-1-4939-7098-8_7.
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Cournac, Axel, Martial Marbouty, Julien Mozziconacci, and Romain Koszul. "Generation and Analysis of Chromosomal Contact Maps of Yeast Species." In Methods in Molecular Biology, 227–45. New York, NY: Springer New York, 2016. http://dx.doi.org/10.1007/978-1-4939-3079-1_13.
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Paro, Renato, Ueli Grossniklaus, Raffaella Santoro, and Anton Wutz. "Genomic Imprinting." In Introduction to Epigenetics, 91–115. Cham: Springer International Publishing, 2021. http://dx.doi.org/10.1007/978-3-030-68670-3_5.
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AbstractA typical cell contains two sets of chromosomes: one that was inherited from the mother, the other from the father. Usually, autosomal alleles are expressed at similar levels from the maternally and paternally inherited chromosomes. This chapter is dedicated to an exception of this rule: the expression of genes that are regulated by genomic imprinting depends on the parental origin of the allele, leading to the non-equivalence of maternal and paternal genomes. Genomic imprinting is a paradigm of epigenetic gene regulation as genetically identical alleles can exist in two expression states within the same nucleus. The imprints marking the parental alleles are established in the parental germline, maintained during the development of the offspring, but reset before they are passed on to the next generation. In mammals, the primary imprint is usually a differentially methylated region at the locus but there are also examples where histone modifications mark the parental alleles. Many imprinted genes play important roles for development and are associated with human disease. Interestingly, genomic imprinting evolved independently in mammals and seed plants and similar mechanisms have been recruited to regulate imprinted expression in the two kingdoms. We will discuss evolutionary constraints that could have led to the evolution of genomic imprinting in these seemingly disparate lineages.
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Kinjo, Y., M. Watanabe, and K. Shinohara. "Comparative Study by Soft X-Ray Contact Microscopy of the Images of Human Chromosomes Treated with Various Conditions." In X-Ray Microscopy III, 442–46. Berlin, Heidelberg: Springer Berlin Heidelberg, 1992. http://dx.doi.org/10.1007/978-3-540-46887-5_100.
Full textConference papers on the topic "Chromosome contacts"
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"Computer tools for spatial chromosome contacts analysis by ChIA-PET and Hi-C data." In SYSTEMS BIOLOGY AND BIOINFORMATICS. Institute of Cytology and Genetics, Siberian Branch of the Russian Academy of Sciences, 2019. http://dx.doi.org/10.18699/sbb-2019-07.
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Silva, Bruno Custódio, Gisele Delazeri, Ana Luíza Kolling Konopka, Giulia Righetti Tuppini Vargas, Paulo Ricardo Gazzola Zen, and Rafael Fabiano Machado Rosa. "Report of a family affected by fragile X syndrome and type 1 diabetes mellitus." In XIII Congresso Paulista de Neurologia. Zeppelini Editorial e Comunicação, 2021. http://dx.doi.org/10.5327/1516-3180.076.
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Context: The fragile X syndrome is characterized by intellectual deficit and some physical characteristics, which become more evident during growth, especially craniofacial and macroorchidism. Case report: A 22 year-old male patient with diabetes mellitus type 1 (DM1) diagnosed at 7 years of age is following-up with ophthalmology due to low visual acuity. On physical exam, he did not maintain eye contact and performed repetitive movements. In addition, he had an elongated face and upward slanting eyelid clefts, a high palate and prognathism, large and prominent ears. In the family history, 3 of his siblings, one male and two female, also had intellectual deficit, and two of them had concomitant DM1. One brother had only DM1 and the other none of the diseases. The parents had consanguinity (they were cousins in the 3rd degree). The patient’s karyotype, using the chromosomal breaks technique after cultivation in medium-low folic acid, showed the presence of fragility on the X chromosome in the region q27.3 [46, XY, fra (x) (q27.3)], compatible with the diagnosis of fragile X syndrome. This result was confirmed using the PCR-multiplex technique. Conclusions: In this family, the concomitant presence in several individuals of the fragile X syndrome and DM1 stands out. However, although both conditions are not related, they are frequent, which could justify their simultaneous occurrence.
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Shinohara, Kunio. "Observation Of Human Chromosomes With Soft X-Ray Contact Microscopy." In 1989 Intl Congress on Optical Science and Engineering, edited by Rene Benattar. SPIE, 1989. http://dx.doi.org/10.1117/12.961823.
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Richardson, M., K. Shinohara, Y. Kinjo, and K. A. Tanaka. "Contact x-ray microscopy of in vitro human chromosome fibers using pulsed laser-plasmas." In OSA Annual Meeting. Washington, D.C.: Optica Publishing Group, 1992. http://dx.doi.org/10.1364/oam.1992.mp2.
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We describe the use of intense pulsed (<1 ns) x- rays from a laser-plasma for making highresolution (100 Å), high-contrast, contact images of hydrated human chromosome fibers. The fibers are suspended in a 10 μm thick H2O cell having a thin 100 nm SiN entrance window anda rear recording panel of PMMA resist. Single-shot irradiation of the cell with x-rays generated from an Au laser-plasma produced by a 5 J, 1 ns duration, laser pulse at 526 nm is sufficient to expose the resist for a cell-target separation of 20 mm. The detailed structure of the image created in three-dimensional relief in the PMMA is faithfully transferred to a thin, positive polymer replica and analyzed with high-resolution electron microscopy. We discuss the implications of this work and describe refinements to this approach that will enhance its usefulness in highresolution imaging of in vitro specimens in the biological and life sciences.
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Shinohara, Kunio, Yasuhito Kinjo, Martin C. Richardson, Atsushi Ito, Noboru Morimoto, Yasuhiro Horiike, Makoto Watanabe, Keiji Yada, and Kazuo A. Tanaka. "Observation of human chromosome fibers in a water layer by laser-plasma x-ray contact microscopy." In San Diego '92, edited by Chris J. Jacobsen and James E. Trebes. SPIE, 1993. http://dx.doi.org/10.1117/12.138754.
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Guttmann, G. D., and J. A. Cobble. "Soft x-ray imaging of biological materials at Los Alamos National Laboratory." In OSA Annual Meeting. Washington, D.C.: Optica Publishing Group, 1992. http://dx.doi.org/10.1364/oam.1992.mp4.
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Soft x-ray contact microscopy (SXCM) is a form of x-ray lithography; however, the biological cell is imaged. Imaging of the biological sample is by soft x- rays within the "water window," which has an energy range of 277–525 eV. The contrast mechanism is based upon the x-ray absorption lengths of carbon and nitrogen, which are ~10 times shorter than that of oxygen in the "water window." In previous experiments at the Lawrence Livermore National Laboratory PHOENIX laser facility, the major result was the first high-resolution (~30 nm) x-ray micrographs of living mammalian cells, CHO-SC1. One of the images showed chromosomes in the mitotic stage. Future efforts at Los Alamos National Laboratory will be the utilization of the Free Electron Laser and in-house laser-produced plasmas as x-ray sources, the use of atomic force microscopy to read the resists, the development of thin window technology and resist applications, the use of multilayer and Fresnel zone plate x-ray optics for holographic purposes, and the use of biological agents to enhance the x-ray images of key cellular structures. Biological studies will look at the structure of chromosomes and chromatin in the living cell.
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Ploos van Amstel, J. K., A. L. van der Zanden, P. H. Reitsma, and R. M. Bertina. "RESTRICTION ANALYSIS AND SOUTHERN BLOTTING OF TOTAL HUMAN DNA REVEALS THE EXISTENCE OF MORE THAN ONE GENE HOMOLOGOUS WITH PROTEIN S cDNA." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1644639.
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A deficiency in protein S, the cofactor of activated protein C, is associated with an increased risk for the development of venous thrombosis. It is inherited as an autosomal dominant disorder. To improve the detection of heterozygotes in affected families, we have started to search for restriction fragment length polymorphism (RFLP) in the protein S gene. This study revealed the existence of two genes containing sequences homologous to protein S cDNA.Three non-overlapping fragments of clone pSUL5, which codes for the carboxy-terminal part of protein S and contains the complete 3' untranslated region, were isolated and used as probes in search for RFLP of the protein S gene.Surprisingly the non-overlapping probes shared more than one hybridizing band. The hybridization took place under stringent assay conditions.This observation is contradictory to the intron-exon organization of a gene and suggests the existence of two genes, containing sequences homologous with pSUL5. Both genes could be assigned to chromosome 3 by mapping through somatic cell hybrids. Whether two functional protein S genes are present in the human genome remains to be established.
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Sadler, J. Evan. "THE MOLECULAR BIOLOGY OF VON WILLEBRAND FACTOR." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643930.
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Human von Willebrand factor (vWF) is a plasma glycoprotein that is synthesized by endothelial cells and megakaryocytes, and perhaps by syncytiotrophoblast of placenta. The biosynthesis of vWF is very complex, involving proteolytic processing, glycosyla-tion, disulfide bond formation, and sulfation. Mature vWF consists of a single subunit of ∼ 250,000 daltons that is assembled into multimer ranging from dimers to species of over 10 million daltons. vWF performs its essential hemostatic function through several binding interactions, forming a bridge between specific receptors on the platelet surface and components of damaged vascular subendothelial connective tissue. Inherited deficiency of vWF, or von Willebrand disease (vWD), is the most common genetically transmitted bleeding disorder worldwide. The last two years has been a time of very rapid progress in understanding the molecular biology of vWF. Four research groups have independently isolated and sequenced the 9 kilobase full-length vWF cDNA. The predicted protein sequence has provided a foundation for understanding the biosynthetic processing of vWF, and has clarified the relationship between vWF and a 75-100 kilodalton plasma protein of unknown function, von Willebrand antigen II (vWAgll)/ vWAgll is co-distributed with vWF in endothelial cells and platelets, and is deficient in patients with vWD. The cDNA sequence of vWF shows that vWAgll is a rather large pro-peptide for vWF, explaining the biochemical and genetic association between the two proteins. vWF has a complex evolutionary history marked by many separate gene segment duplications. The primary structure of the protein contains four distinct types of repeated domains present in two to four copies each. Repeated domains account for over 90 percent of the protein sequence. This sequence provides a framework for ordering the functional domains that have been defined by protein chemistry methods. A tryptic peptide from the amino-terminus of vWF that overlaps domain D3 binds to factor VIII and also appears to bind to heparin. Peptides that include domain A1 bind to collagens, to heparin, and to platelet glycoprotein Ib. A second collagen binding site appears to lie within domain A3. The vWF cDNA has been expressed in heterologous cells to produce small amounts of functionally and structurally normal vWF, indicating that endothelial cells are not unique in their ability to process and assemble vWF multimers. Site-directed mutagenesis has been used to show that deletion of the propeptide of vWF prevents the formation of multimers. Cloned cDNA probes have been employed to isolate vWF genomic DNA from cosmid and λ-phage libraries, and the size of the vWF gene appears to be ∼ 150 kilobases. The vWF locus has been localized to human chromosome 12p12—pter. Several intragenic RFLPs have been characterized. With them, vWF has been placed on the human genetic linkage map as the most telomeric marker currently available for the short arm of chromosome 12. A second apparently homologous locus has been identified on chromosome 22, but the relationship of this locus to the authentic vWF gene is not yet known. The mechanism of vWD has been studied by Southern blotting of genomic DNA with cDNA probes in a few patients. Three unrelated pedigrees have been shown to have total deletions of the vWF gene as the cause of severe vWD (type III). This form of gene deletion appears to predispose to the development of inhibitory alloantibodies to vWF during therapy with cryoprecipitate. During the next several years recombinant DNA methods will continue to contribute our understanding of the evolution, biosynthesis, and structure-function relationships of vWF, as well as the mechanism of additional variants of vWD at the level of gene structure.
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Dousseau, Gabriella Corrêa, Julian Letícia de Freitas, Luíza Alves Corazza, Cristiane de Araújo Martins Moreno, and Maria Sheila Guimarães Rocha. "Arginase 1 deficiency: a differential for progressive ataxia and intellectual disability." In XIV Congresso Paulista de Neurologia. Zeppelini Editorial e Comunicação, 2023. http://dx.doi.org/10.5327/1516-3180.141s1.466.
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Introduction: Arginase-1 deficiency (OMIM #207800) is a rare autosomal recessive genetic disorder caused by mutations in the ARG1 gene, resulting in partial or complete loss of enzyme function that affects the liver-based urea cycle. Spastic paraparesis with onset in early childhood is the most obvious sign of the disease but other symptoms include irritability, delayed growth and development, ataxia, recurrent vomiting, feeding/protein aversion, and anorexia. Brain imaging may reveal cerebral atrophy and cerebellar atrophy may also occur. The ARG1 gene sits on chromosome 6 (6q23) and there are at least 43 potentially disease-causing variants in ARG1. Case report: A 56-year-old female patient with no parental consanguinity and unknown perinatal history — she had no contact with her parents. Since childhood, she reports never liked meat and that she had a lack of appetite and frequent vomiting. She also reports irritability without triggers and difficulties at school — illiterate despite 4 years of schooling. She denied gait difficulties until it was around 50 years, when she presented with progressive imbalance and incoordination. She is currently unable to walk without bilateral support due to severe gait ataxia in addition to appendicular ataxia. Results: Extensive laboratory metabolic investigation was performed, serologies, electroencephalogram and cerebrospinal fluid without alterations. Brain magnetic resonance imaging showed diffuse cortical and cerebellar atrophy. Whole-exome sequencing revealed a homozygous pathogenic variant mutation p.Arg308Gln in the ARG1 gene. Conclusion: In systematic reviews, spastic paraparesis is the hallmark of the disease caused by ARG1 deficiency, however, there are descriptions of rare, predominantly ataxic cases. In basically everyone, intellectual impairment is expected. Therefore, ARG1 deficiency is still under construction and can also be a differential diagnosis for ataxia and intellectual impairment.
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Huber, P., J. Dalmon, M. Laurent, G. Courtois, D. Thevenon, and G. Marguerie. "CHARACTERIZATION OFTHE 5’FLANKING REGION FOR THE HUMAN FIBRINOGEN β GENE." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1642889.
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Fibrinogen is coded by three separate genes located in a 50kb region of chromosome 4 and organized in a α - β - γ orientation with an inversion of the gene 3- A human genomic library was constructed using the EMBL4 phage and screened with cDNA probes coding for human fibrinogen Aα, Bβ and γ chains. Clones, covering the fibrinogen locus,were identified, and their organization was analyzed by means of hybridization and restriction mapping. Among these clones one recombinant phage containing the β gene and large 5’ and 3’ -flanking sequences was isolated.To identify the regulatory sequences Dpstream from the human β gene, a 1.5 kb fragment of the immediate 5’-flanking region was sequenced. The SI mapping experiments revealed three transcription initiation sites. PotentialTATA and CAAT sequences were identified upstream the initiation start points at the positions -21 and -58 from the first initiation start point.Comparison of this sequence with that previously reported for the same region upstream from the human γ gene revealed no significant homology which suggests that the potential promoting sequences of these genes are different. In contrast, comparison of the 5’flanking regions of human and rat β genes showed more than 80% homology for 142 bp upstream from the gene. This highly conserved region is a potential candidate for a regulatory sequence of the human β gene.To verify this activity, a β fibrinogen minigene was constructed by deletion of the internal part of the normal gene and including 3.4kb of the 5’flanking region and 1.4kb of the 3’flanking region. The minigene was transfected into HepG2, a human hepatoma cell line, to show whether the 5’flanking region of the human fibrinogen gene contains DNA sequences sufficient for efficient transcription in HepG2. Constructions of several parts of the sequenced 5’flanking region of the human β gene with the gene of the chloramphenical acetyl transferase have been also transfected in the HepG2 cells to determine the specificity of the gene expression and to localize the sequences controlling the transcription of the gene.
Reports on the topic "Chromosome contacts"
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Lindow, Steven, Isaac Barash, and Shulamit Manulis. Relationship of Genes Conferring Epiphytic Fitness and Internal Multiplication in Plants in Erwinia herbicola. United States Department of Agriculture, July 2000. http://dx.doi.org/10.32747/2000.7573065.bard.
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Most bacterial plant pathogens colonize the surface of healthy plants as epiphytes before colonizing internally and initiating disease. The epiphytic phase of these pathogens is thus an important aspect of their epidemiology and a stage at which chemical and biological control is aimed. However, little is known of the genes and phenotypes that contribute to the ability of bacteria to grow on leaves and survive the variable physical environment in this habitat. In addition, while genes such as hrp awr and others which confer pathogenicity and in planta growth ability have been described, their contribution to other aspects of bacterial epidemiology such as epiphytic fitness have not been addressed. We hypothesized that bacterial genes conferring virulence or pathogenicity to plants also contribute to the epiphytic fitness of these bacteria and that many of these genes are preferentially located on plasmids. We addressed these hypotheses by independently identifying genes that contribute to epiphytic fitness, in planta growth, virulence and pathogenicity in the phytopathogenic bacterium Erwinia herbicola pv gypsophilae which causes gall formation on gypsophila. This species is highly epiphytically fit and has acquired a plasmid (pPATH) that contains numerous pathogenicity and virulence determinants, which we have found to also contribute to epiphytic fitness. We performed saturation transposon mutagenesis on pPATH as well as of the chromosome of E.h. gypsophilae, and identified mutants with reduced ability to grow in plants and/or cause disease symptoms, and through a novel competition assay, identified mutants less able to grow or survive on leaves. The number and identity of plasmid-borne hrp genes required for virulence was determined from an analysis of pPATH mutants, and the functional role of these genes in virulence was demonstrated. Likewise, other pPATH-encoded genes involved in IAA and cytokinin biosynthesis were characterized and their pattern of transcriptional activity was determined in planta. In both cases these genes involved in virulence were found to be induced in plant apoplasts. About half of avirulent mutants in pPATH were also epiphytically unfit whereas only about 10% of chromosomal mutants that were avirulent also had reduced epiphytic fitness. About 18% of random mutants in pPATH were avirulent in contrast to only 2.5% of random chromosomal mutants. Importantly, as many as 28% of pPATH mutants had lower epiphytic fitness while only about 10% of random chromosomal mutants had lower epiphytic fitness. These results support both of our original hypotheses, and indicate that genes important in a variety of interactions with plant have been enriched on mobile plasmids such as pPATH. The results also suggest that the ability of bacteria to colonize the surface of plants and to initiate infections in the interior of plants involves many of the same traits. These traits also appear to be under strong regulatory control, being expressed in response to the plant environment in many cases. It may be possible to alter the pattern of expression of such genes by altering the chemical environment of plants either by genetic means or by additional or chemical antagonists of the plant signals. The many novel bacterial genes identified in this study that are involved in plant interactions should be useful in further understanding of bacterial plant interactions.
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Manulis-Sasson, Shulamit, Christine D. Smart, Isaac Barash, Laura Chalupowicz, Guido Sessa, and Thomas J. Burr. Clavibacter michiganensis subsp. michiganensis-tomato interactions: expression and function of virulence factors, plant defense responses and pathogen movement. United States Department of Agriculture, February 2015. http://dx.doi.org/10.32747/2015.7594405.bard.
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Clavibactermichiganensissubsp. michiganensis(Cmm), the causal agent of bacterial wilt and canker of tomato, is the most destructive bacterial disease of tomato causing substantial economic losses in Israel, the U.S.A. and worldwide. The goal of the project was to unravel the molecular strategies that allow Cmm, a Gram-positive bacterium, to develop a successful infection in tomato. The genome of Cmm contains numerous genes encoding for extracellular serine proteases and cell wall degrading enzymes. The first objective was to elucidate the role of secreted serine proteases in Cmm virulence. Mutants of nine genes encoding serine proteases of 3 different families were tested for their ability to induce wilting, when tomato stems were puncture-inoculated, as compared to blisters formation on leaves, when plants were spray-inoculated. All the mutants showed reduction in wilting and blister formation as compared to the wild type. The chpCmutant displayed the highest reduction, implicating its major role in symptom development. Five mutants of cell wall degrading enzymes and additional genes (i.e. perforin and sortase) caused wilting but were impaired in their ability to form blisters on leaves. These results suggest that Cmm differentially expressed virulence genes according to the site of penetration. Furthermore, we isolated and characterized two Cmmtranscriptional activators, Vatr1 and Vatr2 that regulate the expression of virulence factors, membrane and secreted proteins. The second objective was to determine the effect of bacterial virulence genes on movement of Cmm in tomato plants and identify the routes by which the pathogen contaminates seeds. Using a GFP-labeledCmm we could demonstrate that Cmm extensively colonizes the lumen of xylem vessels and preferentially attaches to spiral secondary wall thickening of the protoxylem and formed biofilm-like structures composed of large bacterial aggregates. Our findings suggest that virulence factors located on the chp/tomAPAI or the plasmids are required for effective movement of the pathogen in tomato and for the formation of cellular aggregates. We constructed a transposon plasmid that can be stably integrated into Cmm chromosome and express GFP, in order to follow movement to the seeds. Field strains from New York that were stably transformed with this construct, could not only access seeds systemically through the xylem, but also externally through tomato fruit lesions, which harbored high intra-and intercellular populations. Active movement and expansion of bacteria into the fruit mesocarp and nearby xylem vessels followed, once the fruit began to ripen. These results highlight the ability of Cmm to invade tomato fruit and seed through multiple entry routes. The third objective was to assess correlation between disease severity and expression levels of Cmm virulence genes and tomato defense genes. The effect of plant age on expression of tomato defense related proteins during Cmm infection was analyzed by qRT-PCR. Five genes out of eleven showed high induction at early stages of infection of plants with 19/20 leaves compared to young plants bearing 7/8 leaves. Previous results showed that Cmm virulence genes were expressed at early stages of infection in young plants compared to older plants. Results of this study suggest that Cmm virulence genes may suppress expression of tomato defense-related genes in young plants allowing effective disease development. The possibility that chpCis involved in suppression of tomato defense genes is currently under investigation by measuring the transcript level of several PR proteins, detected previously in our proteomics study. The fourth objective was to define genome location and stability of virulence genes in Cmm strains. New York isolates were compared to Israeli, Serbian, and NCPPB382 strains. The plasmid profiles of New York isolates were diverse and differed from both Israeli and Serbian strains. PCR analysis indicated that the presence of putative pathogenicity genes varied between isolates and highlighted the ephemeral nature of pathogenicity genes in field populations of Cmm. Results of this project significantly contributed to the understanding of Cmm virulence, its movement within tomato xylem or externally into the seeds, the role of serine proteases in disease development and initiated research on global regulation of Cmm virulence. These results form a basis for developing new strategies to combat wilt and canker disease of tomato.