The radial arrangement of the human chromosome 7 in the lymphocyte cell nucleus is associated with chromosomal band gene density

This is the author's accepted manuscript. The final published article is available from the link below. Copyright @ Springer-Verlag 2008.In the nuclei of human lymphocytes, chromosome territories are distributed according to the average gene density of each chromosome. However, chromosomes are very heterogeneous in size and base composition, and can contain both very gene-dense and very gene-poor regions. Thus, a precise analysis of chromosome organisation in the nuclei should consider also the distribution of DNA belonging to the chromosomal bands in each chromosome. To improve our understanding of the chromatin organisation, we localised chromosome 7 DNA regions, endowed with different gene densities, in the nuclei of human lymphocytes. Our results showed that this chromosome in cell nuclei is arranged radially with the gene-dense/GC-richest regions exposed towards the nuclear interior and the gene-poorest/GC-poorest ones located at the nuclear periphery. Moreover, we found that chromatin fibres from the 7p22.3 and the 7q22.1 bands are not confined to the territory of the bulk of this chromosome, protruding towards the inner part of the nucleus. Overall, our work demonstrates the radial arrangement of the territory of chromosome 7 in the lymphocyte nucleus and confirms that human genes occupy specific radial positions, presumably to enhance intra- and inter-chromosomal interaction among loci displaying a similar expression pattern, and/or similar replication timing

Conservative treatment of a left atrial intramural hematoma after left atrial thrombus resection and concomitant mitral valve replacement - case report

Left atrial intramural hematoma is a seldom cause of left atrial mass. It has been described to occur spontaneously, after interventional procedures, after blunt chest trauma, or after aortocoronary bypass surgery. We present a case of mitral valve replacement together with the removal of a large intraatrial space-occupying lesion. Intraoperative transesophageal echocardiography confirmed a successful resection of this mass. Surprisingly, upon admission to ICU, transesophageal and transthoracic echocardiography revealed a recurrence of an intramural lesion, closest matching a hematoma, which was confirmed by contrast-enhanced computed tomography. Surgical intervention was thoroughly discussed but a conservative management was favoured. 3 months after surgery, a reassessed transthoracic echocardiography and computed tomography demonstrated an almost complete resolution of the pre-existing hematoma

Targeted Deficiency of the Transcriptional Activator Hnf1α Alters Subnuclear Positioning of Its Genomic Targets

DNA binding transcriptional activators play a central role in gene-selective regulation. In part, this is mediated by targeting local covalent modifications of histone tails. Transcriptional regulation has also been associated with the positioning of genes within the nucleus. We have now examined the role of a transcriptional activator in regulating the positioning of target genes. This was carried out with primary β-cells and hepatocytes freshly isolated from mice lacking Hnf1α, an activator encoded by the most frequently mutated gene in human monogenic diabetes (MODY3). We show that in Hnf1a−/− cells inactive endogenous Hnf1α-target genes exhibit increased trimethylated histone H3-Lys27 and reduced methylated H3-Lys4. Inactive Hnf1α-targets in Hnf1a−/− cells are also preferentially located in peripheral subnuclear domains enriched in trimethylated H3-Lys27, whereas active targets in wild-type cells are positioned in more central domains enriched in methylated H3-Lys4 and RNA polymerase II. We demonstrate that this differential positioning involves the decondensation of target chromatin, and show that it is spatially restricted rather than a reflection of non-specific changes in the nuclear organization of Hnf1a-deficient cells. This study, therefore, provides genetic evidence that a single transcriptional activator can influence the subnuclear location of its endogenous genomic targets in primary cells, and links activator-dependent changes in local chromatin structure to the spatial organization of the genome. We have also revealed a defect in subnuclear gene positioning in a model of a human transcription factor disease

Evidence for Sequential and Increasing Activation of Replication Origins along Replication Timing Gradients in the Human Genome

Genome-wide replication timing studies have suggested that mammalian chromosomes consist of megabase-scale domains of coordinated origin firing separated by large originless transition regions. Here, we report a quantitative genome-wide analysis of DNA replication kinetics in several human cell types that contradicts this view. DNA combing in HeLa cells sorted into four temporal compartments of S phase shows that replication origins are spaced at 40 kb intervals and fire as small clusters whose synchrony increases during S phase and that replication fork velocity (mean 0.7 kb/min, maximum 2.0 kb/min) remains constant and narrowly distributed through S phase. However, multi-scale analysis of a genome-wide replication timing profile shows a broad distribution of replication timing gradients with practically no regions larger than 100 kb replicating at less than 2 kb/min. Therefore, HeLa cells lack large regions of unidirectional fork progression. Temporal transition regions are replicated by sequential activation of origins at a rate that increases during S phase and replication timing gradients are set by the delay and the spacing between successive origin firings rather than by the velocity of single forks. Activation of internal origins in a specific temporal transition region is directly demonstrated by DNA combing of the IGH locus in HeLa cells. Analysis of published origin maps in HeLa cells and published replication timing and DNA combing data in several other cell types corroborate these findings, with the interesting exception of embryonic stem cells where regions of unidirectional fork progression seem more abundant. These results can be explained if origins fire independently of each other but under the control of long-range chromatin structure, or if replication forks progressing from early origins stimulate initiation in nearby unreplicated DNA. These findings shed a new light on the replication timing program of mammalian genomes and provide a general model for their replication kinetics

Stable chromosomal units determine the spatial and temporal organization of DNA replication

DNA replication occurs in mammalian cells at so-called replication foci occupying defined nuclear sites at specific times during S phase. It is an unresolved problem how this specific spatiotemporal organization of replication foci is determined. Another unresolved question remains as to what extent DNA is redistributed during S phase. To investigate these problems, we visualized the replicating DNA and the replication machinery simultaneously in living HeLa cells. Time-lapse analyses revealed that DNA was not redistributed to other nuclear sites during S phase. Furthermore, the results showed that DNA is organized into stable aggregates equivalent to replication foci. These aggregates, which we call sub-chromosomal foci, stably maintained their replication timing from S phase to S phase. During S-phase progression, the replication machinery sequentially proceeded through spatially adjacent sets of sub-chromosomal foci. These findings imply that the specific nuclear substructure of chromosomes and the order of their stable subunits determine the spatiotemporal organization of DNA replication

The nuclear organization of Polycomb/Trithorax group response elements in larval tissues of Drosophila melanogaster.

We analysed the nuclear organization of the Polycomb/Trithorax group response element (PRE/TRE) Fab-7 and of other PRE/TREs in larval tissues of D. melanogaster. The results show that pairing/clustering of transgenic and endogenous Fab-7 elements and of other endogenous PRE/TREs occurs only to a limited degree in a highly locus-specific and tissue-specific manner. However, transgenic Fab-7 elements as well as the Fab-7-regulated Abd-B gene and other endogenous loci preferentially occupied defined nuclear regions. Preferred association with the nuclear periphery was observed in the inactive state. However, also in the active state, Fab-7 was often found associated with the nuclear periphery as well as with the boundary of heterochromatin in a fly line- and tissue-specific manner. The boundary between heterochromatin and euchromatin revealed a highly complex architecture in the three-dimensional nuclear space with a close juxtaposition of active and repressed domains. The results suggest that such complex architectures create nuclear microenvironments sustaining specific states of activity of defined PRE/TREs. However, the data also show that the positional behaviour of the transgenic Fab-7 element does not apply to PRE/TREs in general. Altogether, this finding and the highly locus-, tissue-, and fly line-specific behaviour with regard to nuclear positioning and pairing/clustering suggest that the relationships between nuclear organization and functional regulation of PRE/TREs are highly complex and that simple models making general predictions might not be appropriate

The Expression of the Senescence-Associated Biomarker Lamin B1 in Human Breast Cancer

Senescence is a major response to cancer chemotherapy and has been linked to unfavorable therapy outcomes. Lamin B1 is a component of the nuclear lamina that plays a pivotal role in chromatin stability. Downregulation of lamin B1 represents an established biomarker for cellular senescence. However, the protein expression level of lamin B1 in malignant tissue, particularly of the breast, has not been previously described. In this work, we investigated lamin B1 protein expression in normal breast epithelium, malignant breast tissue (including adjacent non-malignant tissue) and in malignant tissue exposed to neoadjuvant chemotherapy (NAC) using immunohistochemistry (IHC) in three patient groups (n = 15, n = 87, and n = 43, respectively). Our results indicate that lamin B1 mean positive expression was 93% in normal breast epithelium and 88% in malignant breast cells, but significantly decreased (mean: 55%, p &lt; 0.001) in malignant breast tissue after exposure to NAC, suggestive of senescence induction. No significant association between lamin B1 expression and other clinicopathological characteristics or survival of breast cancer patients was recorded. To our knowledge, this is the first report that established the baseline protein expression level of lamin B1 in normal and malignant breast tissue, and its reduction following exposure to chemotherapy. In conclusion, lamin B1 downregulation can be used reliably as a component of multiple biomarker batteries to identify therapy-induced senescence (TIS) in clinical cancer.</jats:p