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The occurrence and formation of genomic structural variants (SVs) is known to be influenced by the 3D chromatin architecture, but the extent and magnitude have been challenging to study. Here, we apply Hi-C to study chromatin organization before and after induction of chromothripsis in human cells. We use Hi-C to manually assemble the derivative chromosomes following the occurrence of massive complex rearrangements, which allows us to study the sources of SV formation and their consequences on gene regulation. We observe an action–reaction interplay whereby the 3D chromatin architecture directly impacts the location and formation of SVs. In turn, the SVs reshape the chromatin organization to alter the local topologies, replication timing, and gene regulation in cis. We show that SVs have a strong tendency to occur between similar chromatin compartments and replication timing regions. Moreover, we find that SVs frequently occur at 3D loop anchors, that SVs can cause a switch in chromatin compartments and replication timing, and that this is a major source of SV-mediated effects on nearby gene expression changes. Finally, we provide evidence for a general mechanistic bias of the 3D chromatin on SV occurrence using data from more than 2700 patient-derived cancer genomes.

Warming climate and increasing desertification urge the identification of genes involved in heat and dehydration tolerance to better inform and target biodiversity conservation efforts. Comparisons among extant desert-adapted species can highlight parallel or convergent patterns of genome evolution through the identification of shared signatures of selection. We generate a chromosome-level genome assembly for the canyon mouse (Peromyscus crinitus) and test for a signature of parallel evolution by comparing signatures of selective sweeps across population-level genomic resequencing data from another congeneric desert specialist (Peromyscus eremicus) and a widely distributed habitat generalist (Peromyscus maniculatus), that may be locally adapted to arid conditions. We identify few shared candidate loci involved in desert adaptation and do not find support for a shared pattern of parallel evolution. Instead, we hypothesize divergent molecular mechanisms of desert adaptation among deer mice, potentially tied to species-specific historical demography, which may limit or enhance adaptation. We identify a number of candidate loci experiencing selective sweeps in the P. crinitus genome that are implicated in osmoregulation (Trypsin, Prostasin) and metabolic tuning (Kallikrein, eIF2-alpha kinase GCN2, APPL1/2), which may be important for accommodating hot and dry environmental conditions.

An analysis of competitiveness of thyratrons in relation to up-to-date solid-state devices has been carried out. The design features and parameters of a number of new TGI thyratrons with a hot combined impregnated cathode for operating voltages of 30, 50, and 75 kV, average switching power from 150 to 500 kW and pulsed power to 250 MW, are presented. Thyratrons have a tetrode design with air cooling in the metal ceramic version. The main field of application is high-power electrophysical equipment. In particular, we present the examples of using thyratrons in accelerators as modulators, as well as pulse corona power supplies for air purification systems from hydrogen sulfide, ammonia, and other harmful gases in the streamer corona discharge.

Phenotypic plasticity enables an immediate response to changing conditions, but for most species, evolutionary change through adaptation will be more important for long-term survival. Warming climate and increasing desertification urges the identification of genes involved in heat-and dehydration-tolerance to better inform and target biodiversity conservation efforts. Comparisons among extant desert adapted species can highlight parallel or convergent patterns of genome evolution through the identification of shared signatures of selection. We generate chromosome-level genome assembly for the canyon mouse (Peromyscus crinitus) and test for signature of parallel evolution by comparing signatures of selective sweeps across population-level genomic resequencing data from another desert specialist deer mouse (P. eremicus) and a widely-distributed habitat generalist (P. maniculatus), that may locally adapted to arid conditions. We identify few shared candidate loci involved in desert adaptation and do not find support for a shared pattern of parallel evolution. Instead, we hypothesize divergent molecular mechanisms of desert adaptation among deer mice, potentially tied to species-specific historical demography, which may limit or enhance adaptation. We identify a number of candidate loci experiencing selective sweeps in theP. crinitusgenome that are implicated in osmoregulation (Trypsin, Prostasin) and metabolic regulation (Kallikrein, eIF2-alpha kinase GCN2, APPL1/2), which may be important to accommodating hot and dry environmental conditions.

Higher-order chromatin structure and DNA methylation are implicated in multiple developmental processes, but their relationship to cell state is unknown. Here, we find that large (>7.3 kb) DNA methylation nadirs (termed "grand canyons") can form long loops connecting anchor loci that may be dozens of megabases (Mb) apart, as well as inter-chromosomal links. The interacting loci cover a total of ∼3.5 Mb of the human genome. The strongest interactions are associated with repressive marks made by the Polycomb complex and are diminished upon EZH2 inhibitor treatment. The data are suggestive of the formation of these loops by interactions between repressive elements in the loci, forming a genomic subcompartment, rather than by cohesion/CTCF-mediated extrusion. Interestingly, unlike previously characterized subcompartments, these interactions are present only in particular cell types, such as stem and progenitor cells. Our work reveals that H3K27me3-marked large DNA methylation grand canyons represent a set of very-long-range loops associated with cellular identity.

Hi-Culfite, a protocol combining Hi-C and whole-genome bisulfite sequencing (WGBS), determines chromatin contacts and DNA methylation simultaneously. Hi-Culfite also reveals relationships that cannot be seen when the two assays are performed separately. For instance, we show that loci associated with open chromatin exhibit context-sensitive methylation: when their spatial neighbors lie in closed chromatin, they are much more likely to be methylated.

Higher order chromatin structure and DNA methylation are implicated in multiple developmental processes, but their relationship to cell state is unknown. Here, we found that large (~10kb) DNA methylation nadirs can form long loops connecting anchor loci that may be dozens of megabases apart, as well as interchromosomal links. The interacting loci comprise ~3.5Mb of the human genome. The data are more consistent with the formation of these loops by phase separation of the interacting loci to form a genomic subcompartment, rather than with CTCF-mediated extrusion. Interestingly, unlike previously characterized genomic subcompartments, this subcompartment is only present in particular cell types, such as stem and progenitor cells. Further, we identify one particular loop anchor that is functionally associated with maintenance of the hematopoietic stem cell state. Our work reveals that H3K27me3-marked large DNA methylation nadirs represent a novel set of very long-range loops and links associated with cellular identity.SummaryHi-C and DNA methylation analyses reveal novel chromatin loops between distant sites implicated in stem and progenitor cell function.

The human genome folds to create thousands of intervals, called “contact domains,” that exhibit enhanced contact frequency within themselves. “Loop domains” form because of tethering between two loci—almost always bound by CTCF and cohesin—lying on the same chromosome. “Compartment domains” form when genomic intervals with similar histone marks co-segregate. Here, we explore the effects of degrading cohesin. All loop domains are eliminated, but neither compartment domains nor histone marks are affected. Loss of loop domains does not lead to widespread ectopic gene activation but does affect a significant minority of active genes. In particular, cohesin loss causes superenhancers to co-localize, forming hundreds of links within and across chromosomes and affecting the regulation of nearby genes. We then restore cohesin and monitor the re-formation of each loop. Although re-formation rates vary greatly, many megabase-sized loops recovered in under an hour, consistent with a model where loop extrusion is rapid. Mapping the nucleome in 4D during cohesin loss and recovery reveals that cohesin degradation eliminates loop domains but has only modest transcriptional consequences. Keywords: cohesion; genome architecture; loop extrusion; chromatin loops; superenhancers; gene regulation; nuclear compartments; Hi-C; 4D Nucleome; CTCF

SUMMARYThe human genome folds to create thousands of intervals, called “contact domains,” that exhibit enhanced contact frequency within themselves. “Loop domains” form because of tethering between two loci - almost always bound by CTCF and cohesin – lying on the same chromosome. “Compartment domains” form when genomic intervals with similar histone marks co-segregate. Here, we explore the effects of degrading cohesin. All loop domains are eliminated, but neither compartment domains nor histone marks are affected. Loci in different compartments that had been in the same loop domain become more segregated. Loss of loop domains does not lead to widespread ectopic gene activation, but does affect a significant minority of active genes. In particular, cohesin loss causes superenhancers to co-localize, forming hundreds of links within and across chromosomes, and affecting the regulation of nearby genes. Cohesin restoration quickly reverses these effects, consistent with a model where loop extrusion is rapid.

Higher order chromatin structure and DNA methylation are implicated in multiple developmental processes, but their relationship to cell state is unknown. In order to understand how the DNA methylation is connected with nuclear architecture and can vary between cell types and during cell differentiation, we began to explore the 3D architecture of human hematopoietic stem and progenitor cells (HSPCs) by performing in situ Hi-C experiments at 5kb resolution. We found that large (~10kb) DNA methylation canyons can form long loops connecting anchor loci that may be dozens of megabases apart. These canyons also can form interchromosomal links (Fig.1a and 1b). We further confirmed these long-range interactions by performing 3D-FISH using two color fluorescent labeled probes that spanned the HOXA locus loop anchor (green) and the SP8 locus loop anchor (red), which are ~7MB apart (Fig. 1c). In order to begin to investigate mechanisms that may regulate these long loops and how they relate to commonly studied loops that are mediated by CTCF-extrusion, we examined their properties systematically. Interestingly, the anchors of long loops exhibited minimal enrichment for CTCF (1.04-fold), and, even when CTCF was bound, they did not obey the convergent rule. The data suggest these loops are formed by phase separation of the interacting loci to form a genomic subcompartment, rather than by CTCF-mediated extrusion. Next, we sought to determine whether other features correlated with these long loops. By aligning DNA methylation profiles with the Hi-C data, we observed that anchors often corresponded to regions of very low DNA methylation, and thus sought to analyze the relationship in detail. We found that the anchor position of the long loops had lower average DNA methylation levels than standard loop anchors and very often overlapped with DNA methylation canyons. Canyons are typically decorated with either active or repressive histone marks. We considered whether a particular group of canyons was associated with the long loops. Our findings further indicate that repressed regions marked by the polycomb-mediated histone modification H3K27me3 at DNA methylation canyons generally mediate the formation of canyon loops. Next, we considered whether the long loops associated with repressive grand canyons that we had annotated in HSPCs were present in other cell types. Using Aggregate Peak Analysis (APA), a computational strategy in which the Hi-C submatrices from the vicinity of multiple putative loops are superimposed, we examined 19 human cell types and 10 murine cell types in which loop-resolution Hi-C maps are available. Interestingly, unlike previously characterized genomic subcompartments, these long-range loops are only present in stem and progenitor cells, but not in differentiated cell types, such as T cells and erythroid progenitors (Fig. 1d). Further, we identified one particular loop anchor that lay at the anchor of a long loop and contained no apparent genes ("geneless" canyon, or "GLS"). The GLS harboring this anchor is 17 kb long, lies 1.4 Mb upstream of the HOXA1 gene, and forms long loops with a 28 kb grand canyon in the HOXA region. In order to understand the role of the GLS region in hematopoietic stem cells (HSCs), we deleted the GLS in HSPCs using Cas9-mediated editing and assayed the edited cells for their ability to form colonies. Strikingly, after deleting the GLS, the number of colonies and their size was greatly reduced in edited cells compared to control experiments using either random guide RNAs or electroporation only (Fig. 1e). After ex vivo culture, the overwhelming majority of GLS-knock out HSPCs acquired the marker CD38, indicating that they were differentiating. Similarly, HOXA gene expression, an indicator of HSPC function, was greatly diminished after GLS deletion compared to control cells. These data indicate that the GLS identified in our study is functionally associated with maintenance of the HSC state. Overall, our work reveals long-range interactions between H3K27me3-marked DNA methylation canyons comprising a novel microcompartment associated with cellular identity. Figure 1. Figure 1. Disclosures No relevant conflicts of interest to declare.

We recently used in situ Hi-C to create kilobase-resolution 3D maps of mammalian genomes. Here, we combine these maps with new Hi-C, microscopy, and genome-editing experiments to study the physical structure of chromatin fibers, domains, and loops. We find that the observed contact domains are inconsistent with the equilibrium state for an ordinary condensed polymer. Combining Hi-C data and novel mathematical theorems, we show that contact domains are also not consistent with a fractal globule. Instead, we use physical simulations to study two models of genome folding. In one, intermonomer attraction during polymer condensation leads to formation of an anisotropic "tension globule." In the other, CCCTC-binding factor (CTCF) and cohesin act together to extrude unknotted loops during interphase. Both models are consistent with the observed contact domains and with the observation that contact domains tend to form inside loops. However, the extrusion model explains a far wider array of observations, such as why loops tend not to overlap and why the CTCF-binding motifs at pairs of loop anchors lie in the convergent orientation. Finally, we perform 13 genome-editing experiments examining the effect of altering CTCF-binding sites on chromatin folding. The convergent rule correctly predicts the affected loops in every case. Moreover, the extrusion model accurately predicts in silico the 3D maps resulting from each experiment using only the location of CTCF-binding sites in the WT. Thus, we show that it is possible to disrupt, restore, and move loops and domains using targeted mutations as small as a single base pair.

The problem of plasma structure stabilization recently has received considerable attention for low and high temperature plasma applications, in particular in fusion experiments, and in order to cope with the problem different special configurations of current and magnetic systems are developed and deployed. However, the available solutions require construction of complex magnetic fields and consume considerable amounts of energy. With respect to thyratrons, which operation is characterized by the presence of normally several high current plasma channels, this issue has not been yet studied thoroughly. Based on experimental data, we analyzed the results of the influence of an external conductive shield on stabilization of plasma channels in high-power pseudospark switches with reentrant and classic design. Both non-ferrous and ferrous shields have been tested. The preliminary calculation of the magnetic field distribution is presented. This research is a part of a work on improvement of switching capabilities of thyratrons used for transferring currents up to hundreds kA with switching energy more than 50 kJ.

In the paper the most recent research and development efforts made by Pulsed Technologies ltd. are presented. Design and essential characteristics of power high-voltage TDI and TPI-thyratrons (Pseudospark Switches), spark gaps, as well as X-ray tubes of the new generation are described. The report cites examples of emerging applications in leading world institutions, including high-power pulsed Electro-Physics (EP).

A pulse power system of the induction electron accelerator intended for parameters of 2 MeV, 2 kA, and operating in the double-pulse mode is presented. The schematic diagram and main elements of the pulse system are described, and the main technical solutions intended to produce a set of pulses with 21-kV peak voltages, peak currents of up to 8 kA, and 200-ns durations across a resistive-inductive load are indicated. The experimental data, which were obtained when separate units and the whole power system operated on the rated duty, are given.

We recently used in situ Hi-C to create kilobase-resolution 3D maps of mammalian genomes. Here, we combine these maps with new Hi-C, microscopy, and genome-editing experiments to study the physical structure of chromatin fibers, domains, and loops. We find that the observed contact domains are inconsistent with the equilibrium state for an ordinary condensed polymer. Combining Hi-C data and novel mathematical theorems, we show that contact domains are also not consistent with a fractal globule. Instead, we use physical simulations to study two models of genome folding. In one, intermonomer attraction during polymer condensation leads to formation of an anisotropic "tension globule." In the other, CCCTC-binding factor (CTCF) and cohesin act together to extrude unknotted loops during interphase. Both models are consistent with the observed contact domains and with the observation that contact domains tend to form inside loops. However, the extrusion model explains a far wider array of observations, such as why loops tend not to overlap and why the CTCF-binding motifs at pairs of loop anchors lie in the convergent orientation. Finally, we perform 13 genome-editing experiments examining the effect of altering CTCF-binding sites on chromatin folding. The convergent rule correctly predicts the affected loops in every case. Moreover, the extrusion model accurately predicts in silico the 3D maps resulting from each experiment using only the location of CTCF-binding sites in the WT. Thus, we show that it is possible to disrupt, restore, and move loops and domains using targeted mutations as small as a single base pair.

A double pulse modulator is described that is capable of generating two pulses of 200 ns duration at peak currents up to 10 kA with the delay between pulses as short as 2 μs. The TPI1-10k/50 tetrode and the TPI5-10k/50 pentode cold cathode thyratrons, which can be operated at voltages up to 50 kV, have been used as switching elements. A voltage hold-off recovery time of less than 1 μs has been demonstrated at an operating current of 7 kA and a reverse voltage hold-off of 24 kV. Unlike common gaseous switches, TPI thyratrons can operate for over 20,000 h at hundreds of kHz. These devices are serious competitors to both conventional hydrogen hot cathode thyratrons and to high-power solid-state switches. TPI thyratrons are particularly advantageous in burst mode operation where direct and reverse voltages in the range of tens of kV, currents up to 10 kA, nanosecond jitter and risetimes of 1-5 ns are required.

The circuit of modulator, serving to supply inductive-resistive load in double-pulse mode with currents up to 10 kA and pulse duration of 300 ns, is described. As switching components unheated cathode thyratrons (pseudospark switches) TPI1-10k/50 and TPI5-10k/50 with anode voltage up to 50 kV have been used. The results of tests, confirming possibility of the thyratron reverse dielectric strength recovery within some microseconds after switching of 10 kA forward anode current, are presented.

The paper demonstrates the results of TDI1-200k/25SN-P pseudospark switch (PSS) developed by Russian company "Pulsed Technologies Ltd" application. PSS was used in pulsed power unit intended for electric-discharge fracture of rocks and concrete blocks and splitting off from monolith. The pulsed power unit has a pulse current generator with the capacity of 560 [mu]F, stored energy of up to 63 kJ, operating voltage of up to15 kV, current pulse amplitude of up to 200 kA and pulse duration more than 200 [mu]sec. The study also shows the current waveforms determined in the short-circuit experiment of the pulse current generator and in the experiments of the electric-discharge fragmentation of concrete at the charging voltage of 13 kV. PSS was operated in ringing single-pulse mode with the exceedance of more than two maximum permissible parameters: current pulse amplitude, current pulse duration and maximum pulse energy. Internal electrode erosion of PSS is shown and possible reasons of asymmetric current feed are discussed.

SummaryWe use in situ Hi-C to probe the 3D architecture of genomes, constructing haploid and diploid maps of nine cell types. The densest, in human lymphoblastoid cells, contains 4.9 billion contacts, achieving 1 kb resolution. We find that genomes are partitioned into contact domains (median length, 185 kb), which are associated with distinct patterns of histone marks and segregate into six subcompartments. We identify ∼10,000 loops. These loops frequently link promoters and enhancers, correlate with gene activation, and show conservation across cell types and species. Loop anchors typically occur at domain boundaries and bind CTCF. CTCF sites at loop anchors occur predominantly (>90%) in a convergent orientation, with the asymmetric motifs “facing” one another. The inactive X chromosome splits into two massive domains and contains large loops anchored at CTCF-binding repeats.PaperFlick

The pseudospark (thyratrons TPI- and TDI-type) switch drivers design and applications in various pulsed power systems are presented. Drivers are constructed using modern IGBT transistors and triggered compact spark gaps, and feature both manual and remote digital control (CAN, Ethernet). TDI-thyratrons can serve as a replacement for ignitron, spark gap and triggered vacuum switches. The TDI drivers are made with multiple outputs for triggering up to 4 thyratrons in parallel or with HV isolated output for applications where the igniter and hydrogen reservoir of the thyratron are under high (transient) voltages. TPI-thyratrons were designed recently however, we have extensive experience in application and the methods of triggering/controlling of these switches. The trigger unit is a simple relaxation oscillator with DC and Pulsed Preionization, including single, double and triple pulse triggering techniques. In particular it is shown that when the requirement for a driver is to provide minimum time jitter (less 0.5ns), high anode current pulse stability and extended lifetime, the best option is triple-pulse triggering. Circuit design for trigger drivers BZ-TP/10 for TPI-thyratrons, optimized for the operation of the particle accelerator LIU-2 are described.

The operation of pseudosparks (thyratrons with cold cathodes) of the TDI1-150k/25 type was investigated in the nanosecond time range of control and switching. Their most important characteristics are their reliability of firing—practically in 100% of cases, a jitter time—less than 4 ns, and the possibility of them switching off (interruption of a current) at a particular moment (after the first half of a discharge cycle). Investigations have shown that the TDI1-150k/25 device has all the above characteristics (which meet the current industrial demands) if the proper operational regimes are chosen. We have proven its parameters with a set of four switches working in parallel at the current level of a few hundred kiloamperes at our installations of the Dense Plasma Focus type, having an energy of several kilojoules.

A pseudospark gap is described that is capable of interrupting a current of up to 1 kA and shaping voltage pulses with an amplitude of up to 110 kV and rise time of ∼100 ns. Analytical expressions for calculating the energy released in the device at the switching-off stage and the efficiency of the generator with an inductive energy storage system are obtained. The characteristics of the pseudospark gap and of a similar-design thyratron connected in the same circuit are compared. It is found that the turn-off time of the pseudospark gap is shorter than that of the thyratron by a factor of 2.5.

DNA methylation is a critical regulator of cis-regulatory elements that can impact the distribution of epigenetic regulators and transcription factors. We have mapped DNA methylation changes in human CD34+ cells and downstream progeny to detect changes in DNA methylation with differentiation. Some large regions of low DNA methylation emerge with differentiation, which we call dynamic (d-) canyons. In order to determine whether these correlate with changes in the 3D genome, we generated 6 high-resolution Hi-C contact maps from CD34+CD38- cells, Adult bone marrow CD34+ cells, 7 days cultured CD34+ cells, CD71+CD36+CD235+ Erythroid cells, CD4+ T cells and AML patient blast cells. We generated ~ 1 billion mapped reads in order to create each 3D map of the genome at kilobase resolution. We identified several sites of d-canyons which correlate with changes in Hi-C loops. To study the functional role of d-canyons, we performed CRISPR/CAS9 mediated genome engineering targeting three sites of interest in CD34+ progenitors. We designed sgRNAs to delete several d-canyon regions that exhibited cell-type specific methylation and dynamic H3K27ac marking. First, we targeted a novel putative regulatory region of the RUNX1 gene, a critical master regulator of hematopoiesis that is frequently mutated in human leukemia. From genome-wide DNA methylation profiling, we identified a d-canyon located in the first intron of RUNX1, which overlaped with a H3K27ac peak in human CD34+CD38- hematopoietic progenitor cells. DNA methylation in this region is further depleted in T cells and increased in AML cells, suggesting a role for regulating RUNX1 in specific cell types. To study its function, we designed 2 sgRNAs to delete a 1.8 kb d-canyon. CRISPR/Cas9-mediated deletion of this region resulted in ablation of RUNX1 expression in cord blood hematopoietic stem cells and a significant decrease of engraftment activity in NSG mice along with an increase of erythroid colony forming ability in in vitro assays. In addition, we identified d-canyons upstream of the master regulators GATA2 and in the HOXA cluster. We deleted 1.7kb d-canyon upstream of GATA2 gene, finding that it resulted in increased self-renewal of CD34+CD38- cells in NSG xenografts. In contrast, when we deleted one a d-canyon located ~2Mb upstream of the HOXA cluster, within a HiC loop signal present only in CD34+CD38- cells, we observed in decreased CD34+CD38- cell self-renewal but a significant increase of CD34+CD38+ differentiated progenitors in an in vitro culture system, as well as in NSG mice. Colony forming assay showed decreased colony size and numbers. Finally, we identified a d-canyon associated with TCF3, also known as E2A, a transcription factor involved in B and T cell lineage differentiation. We designed 4 sgRNAs to delete 1.5kb d-canyon edge regions within the second intron of TCF3. Removal of this region resulted in a significant decrease of CD19+ B cells, but an increase of CD3+ T cells in NSG mice. Taken together, these results suggest the functional importance of d-canyons for orchestrating genome architecture and fate decisions of hematopoietic stem cells. These findings advance our understanding of the relationship between DNA methylation changes and loop interactions, providing new insights into the potential impact of potential aberrant DNA methylation and chromatin structure. Disclosures No relevant conflicts of interest to declare.

High order chromatin structure is implicated in multiple developmental processes and disease. However, a global picture of chromosomal looping interaction alterations during stem cell self-renewal and differentiation is lacking. Hematopoietic stem cell (HSCs) and their differentiated progenitors (HSPCs) offer a system in which to examine this. Of the key differentiated lineages, the erythroid lineage undergoes a unique nuclear condensation process during a well-characterized differentiation process which can be induced in vitro from CD34+ HSPCs. Thus erythroid differentiation offers an ideal model system to study differentiation-associated changes in high order chromatin structure. We have thus generated the in situ Hi-C contact map for human cord blood CD34+ CD38- HSPC (CD34+) and erythroid progenitors undergoing differentiation in vitro at day 7 from CD34+ HSPCs (EryD7). In our 5kb resolution map, we identified over 2000 chromosomal loop interactions in both CD34+ and Day 7 erythroid respectively . The EryD7 sample exhibited higher random intra-chromosomal interactions in comparison with CD34+, presumably due to nuclear condensation. By comparing the chromosomal loop interactions in the 2 cell types. We identified self-renewal and erythroid differentiation-specific looping patterns in the two cell types. Strikingly, we found that a gene depleted region (GDR) 2MB upstream of the HOXA cluster forms a strong chromosome loop with the HOXA cluster exclusively in the HSPCs (Fig1A). Within this GDR site, we identified two conserved CTCF sites, which are thought to organized chromosome looping. Utilizing the CRISPR-mediated deletion of each of the two CTCF sites, we found that deletion of either site reduce the colony forming ability of CD34+, indicating a loss of stem cell self-renewal. (Fig 1B) Gene expression analysis showed that HOXA9 expression was compromised the CTCF site deletion. These data suggest that the GDR is forming a distant regulatory loop which controls the expression of HOXA9 in HSPCs. Because the GDR is implicated in controlling HOXA9 expression, a key gene in leukemogenesis, we then tested the importance of this looping site in different leukemia cell lines that are dependent on HOXA9. Of those cell lines, we found the deletion of the CTCF sites inhibit the growth of DNMT3A and NPM1 mutated OCI-AML3 and promote the apoptosis. In contrast, growth of the MLL translocation cell line MV 4:11 was not abrogated by their deletion (Fig 1C). As a control cell line which doesn't express HOXA9, HL60 cells were not sensitive to the deletion of the GDR CTCF sites. Together, these data indicate leukemic cells may adopt different strategies to activate HOXA9. MLL translocation leukemias activates HOXA9 by the direct binding of the MLL fusion protein, while the NPM1 mutated leukemia is more likely to utilize the stem cell looping to activate HOXA9 expression. Among EryD7 specific interactions, we found the β-globin locus specifically forms chromatin loops at Day7 that are not evident in the CD34+ HSPCs. Detailed examination showed that Dnase I hypersensitivity sites HS5 and 3'HS1 both contains CTCF site and form chromosomal loops. Two other loop-forming CTCF sites, both on the telomeric and centromeric side of β-globin locus were also identified. Interestingly, we found a CTCF binding site adjacent to OR52A5 gene which forms a chromosomal loop with HS5 and is not well studied. To test the role of the chromosomal looping in the regulation of hemoglobin gene expression in β-globin locus, we deleted the OR52A5-CTCF site and the 3'HS1 CTCF site in K562 and adult CD34+ HSPCs. We found the deletion of OR52A5-CTCF resulted in a decrease of HBE and increase of HBB expression in K562 cells, which suggest the OR52A5 CTCF also plays a role in regulating hemoglobin gene expression in the β-globin locus (Fig 1D). Furthermore, we found the deletion of 3'HS1 CTCF resulted in a 4-fold increase of HBG2 expression in adult CD34+ HSPC during erythroid differentiation (Fig1 E). Thus this indicating the 3'HS1 and OR52A5-CTCF CTCF sites in β-globin locus are forming loops that regulate the β-globin locus gene expression. In summary, we have mapped the higher order chromatin structure alterations during stem cell differentiation and identified the critical looping interaction essential for the self-renewal and differentiation specific functions. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Methods of measuring charges in the bulk and at the surface of the insulating envelope of a high-voltage vacuum device are developed as part of studies of a complex range of dielectric strength aspects. These methods were used to measure the charge distribution over the length of the envelope and to study how this distribution is influenced by the operating regimes of the device. Laws governing the formation of charges were identified and its correlation with the appearance of physical changes in the structure of the surface layers of the insulator was determined. Processes of structural change were studied and simulated and the change in the dielectric strength of the device was compared with changes in the magnitude and polarity of the surface and bulk charges.

A new atmospheric electromagnetic plasmadynamic system, pulsed-periodic plasmatron, operating at standard atmospheric pressure with high frequency has been developed. In this system plasma flow velocity reaches 3–5·103 m/s with temperature up to 15·103 K. The results are presented on the use of the pulsed-periodic plasmatron for surface layer modification and particles acceleration.

The issues of improvement of hold-off capability and reliability of high-voltage vacuum tubes, for example, X-ray tubes were studied. Two methods are suggested for improvement of the tubes operation. The first method is based on deposition of special coatings on the inner surface of tube isolator in the regions where electric field strength is high. These coatings have bulk electric conductivity higher than for isolator material but their surface conductivity is almost equal. The second method is based on pulsed surface melting of the tube electrodes with non-relativistic, high-current electron beam. Pulsed surface melting drastically reduces parasitic field emission, which results in the decrease of the charge density both in the bulk and surface of isolator providing high hold-off voltages through the tube. The tests of 160-kV and 250-kV X-ray tubes made utilizing the new above mentioned technologies have shown that parasitic currents do not exceed 1 μA and conditioning time decreases from 5 hours to 15 minutes. The new technology also allows to reduce dimensions of the tube.

We present a dynamical model of the high mass X-ray binary LMC X-1 based on high-resolution optical spectroscopy and extensive optical and near-infrared photometry. From our new optical data we find an orbital period of P=3.90917 +/- 0.00005 days. We present a refined analysis of the All Sky Monitor data from RXTE and find an X-ray period of P=3.9094 +/- 0.0008 days, which is consistent with the optical period. A simple model of Thomson scattering in the stellar wind can account for the modulation seen in the X-ray light curves. The V-K color of the star (1.17 +/- 0.05) implies A_V = 2.28 +/- 0.06, which is much larger than previously assumed. For the secondary star, we measure a radius of R_2 = 17.0 +/- 0.8 solar radii and a projected rotational velocity of V_rot*sin(i) = 129.9 +/- 2.2 km/s. Using these measured properties to constrain the dynamical model, we find an inclination of i = 36.38 +/- 1.92 deg, a secondary star mass of M_2 = 31.79 +/- 3.48 solar masses, and a black hole mass of 10.91 +/- 1.41 solar masses. The present location of the secondary star in a temperature-luminosity diagram is consistent with that of a star with an initial mass of 35 solar masses that is 5 Myr past the zero-age main sequence. The star nearly fills its Roche lobe (~90% or more), and owing to the rapid change in radius with time in its present evolutionary state, it will encounter its Roche lobe and begin rapid and possibly unstable mass transfer on a timescale of a few hundred thousand years., Comment: 54 manuscript pages, 14 figures (two of degraded quality), 9 tables, to appear in ApJ. Several revisions to matched the accepted version, but only minor changes in the adopted parameters. The circular orbit model is preferred over the eccentric orbit model

Summary form only given. A modulator circuit providing for supply of inductive-resistive load in double-pulse mode with currents up to 10 kA and pulse duration of 300 ns is described. As switching components thyratrons (pseudospark switches) TPI1-10k/50 with anode voltage up to 50 kV are used. In contrast with other pseudospark switches TPI-thyratrons are capable of operating in circuitry with grounded cathode which allows a service life time up to 20,000 hours. The thyratrons can be considered as an alternative to well-known switches, including hot cathode hydrogen thyratrons and up-to-date power solid state switches, especially for switching of high-currents with sub-nanosecond jitter, turn-on time of 3-5 ns and average current up to 0.3 A. The absence of hot cathode provides in thyratron more homogeneous distribution of gas density and hence shorter turn-on time. Simultaneously it simplifies thyratron design, removes additional power supply and source of heat. There is no need for forced cooling. This new design allows operation without cathode heating and, which is more valuable, without hydrogen reservoir heating. This significantly simplifies auxiliary circuits up to a level of spark gaps. Results of tests, confirming the possibility of reverse dielectric strength recovery within some microseconds after switching of 10 kA forward anode current are presented. Basic electrical parameters affecting thyratron operation in the indicated mode are analyzed. A pentode version of the thyratron, which allows for a reduced recovery time up to 0.5-1 mus is designed and will be presented.

The data on the occurrence of HIV in Russia in 1987-1991 are presented and the effects of the epidemiological surveillance system used at that time on the epidemiological process of HIV infection in the Russian Federation is evaluated. The most important features of that system included the centralized registration of all cases of HIV infection irrespective of the degree of clinical symptoms, large-scale obligatory examination of the population for antibody to HIV, epidemiological investigation of all reported cases of HIV infection. The occurrence of HIV infection in 1987-1991 in Russia remained very low, 0.335 per 100,000, showing no trend for increase. Owing to that system of the epidemiological surveillance of HIV infection in the USSR, nosocomial outbreaks were discovered which gave a rise in the number of recorded cases in 1989. In 1990-1991, changes in the structure of the epidemic process occurred. In those years, HIV transmission among homosexuals became most important, increasing from 22.4% to 47.8% which characterized the beginning of a new stage of the epidemic. On the whole, the epidemiological surveillance system for HIV in the USSR, 1987-1991, was effective, although it needs improvement including reconsideration of the population groups to be obligatorily examined for HIV antibody.