CTCF-mediated transcriptional regulation through cell type-specific chromosome organization in the {\beta}-globin locus

The principles underlying the architectural landscape of chromatin beyond the nucleosome level in living cells remains largely unknown despite its potential to play a role in mammalian gene regulation. We investigated the 3-dimensional folding of a 1 Mbp region of human chromosome 11 containing the {\beta}-globin genes by integrating looping interactions of the insulator protein CTCF determined comprehensively by chromosome conformation capture (3C) into a polymer model of chromatin. We find that CTCF-mediated cell type specific interactions in erythroid cells are organized to favor contacts known to occur in vivo between the {\beta}-globin locus control region (LCR) and genes. In these cells, the modeled {\beta}-globin domain folds into a globule with the LCR and the active globin genes on the periphery. By contrast, in non-erythroid cells, the globule is less compact with few but dominant CTCF interactions driving the genes away from the LCR. This leads to a decrease in contact frequencies that can exceed 1000-fold depending on the stiffness of the chromatin and the exact positioning of the genes. Our findings show that an ensemble of CTCF contacts functionally affects spatial distances between control elements and target genes contributing to chromosomal organization required for transcription.Comment: Full article, including Supp. Mat., is available at Nucleic Acids Research, doi: 10.1093/nar/gks53

The Isl1/Ldb1 complex orchestrates heart-specific chromatin organization and transcriptional regulation

Cardiac stem/progenitor cells hold great potential for regenerative therapies however the mechanisms regulating their expansion and differentiation remain insufficiently defined. Here we show that the multi-adaptor protein Ldb1 is a central regulator of cardiac progenitor cell differentiation and second heart field (SHF) development. Mechanistically, we demonstrate that Ldb1 binds to the key regulator of SHF progenitors Isl1 and protects it from proteasomal degradation. Furthermore, the Isl1/Ldb1 complex promotes long-range promoter-enhancer interactions at the loci of the core cardiac transcription factors Mef2c and Hand2. Chromosome conformation capture followed by sequencing identified surprisingly specific, Ldb1-mediated interactions of the Isl1/Ldb1 responsive Mef2c anterior heart field enhancer with genes which play key roles in cardiac progenitor cell function and cardiovascular development. Importantly, the expression of these genes was downregulated upon Ldb1 depletion and Isl1/Ldb1 haplodeficiency. In conclusion, the Isl1/Ldb1 complex orchestrates a network for heart-specific transcriptional regulation and coordination in three-dimensional space during cardiogenesis

Targeted transgene integration overcomes variability of position effects in zebrafish.

Zebrafish transgenesis is increasingly popular owing to the optical transparency and external development of embryos, which provide a scalable vertebrate model for in vivo experimentation. The ability to express transgenes in a tightly controlled spatio-temporal pattern is an important prerequisite for exploitation of zebrafish in a wide range of biomedical applications. However, conventional transgenesis methods are plagued by position effects: the regulatory environment of genomic integration sites leads to variation of expression patterns of transgenes driven by engineered cis-regulatory modules. This limitation represents a bottleneck when studying the precise function of cis-regulatory modules and their subtle variants or when various effector proteins are to be expressed for labelling and manipulation of defined sets of cells. Here, we provide evidence for the efficient elimination of variability of position effects by developing a PhiC31 integrase-based targeting method. To detect targeted integration events, a simple phenotype scoring of colour change in the lens of larvae is used. We compared PhiC31-based integration and Tol2 transgenesis in the analysis of the activity of a novel conserved enhancer from the developmentally regulated neural-specific esrrga gene. Reporter expression was highly variable among independent lines generated with Tol2, whereas all lines generated with PhiC31 into a single integration site displayed nearly identical, enhancer-specific reporter expression in brain nuclei. Moreover, we demonstrate that a modified integrase system can also be used for the detection of enhancer activity in transient transgenesis. These results demonstrate the power of the PhiC31-based transgene integration for the annotation and fine analysis of transcriptional regulatory elements and it promises to be a generally desirable tool for a range of applications, which rely on highly reproducible patterns of transgene activity in zebrafish.This work was funded by ‘BOLD’ Marie-Curie Initial Training Network; and ‘ZFHealth’ Integrating project in the Framework 7 programme of the European RESEARCH ARTICLE Development (2014) doi:10.1242/dev.100347Development Commission; University of Birmingham (F.M.); Temple University; and the National Institutes of Health (NIH) [HD061749 to D.B.]

Role of LDB1 in the transition from chromatin looping to transcription activation

Şcoala de Management în Sănătate Publică, Universitatea de Stat de Medicină şi Farmacie „Nicolae Testemiţanu”, Chişinău, Republica MoldovaÎncepând cu anul 2001, Ministerul Sănătăţii, factorii de decizie locali şi internaţionali, au iniţiat realizarea obiectivelor de dezvoltare a capacităţilor în Managementul Sănătăţii Publice prin crearea Şcolii de Management în Sănătate Publică din Republica Moldova, care şi-a început activitatea în anul 2003. Prezenta lucrare abordează totalizarea eforturilor de dezvoltare a capacităţilor în domeniul managementului sănătăţii publice pe parcursul ultimului deceniu (2005-2015), în special, prin organizarea programului de masterat în domeniul nominalizat în Republica Moldova. Aspectele elucidate cuprind componentele de instruire, cercetare, servicii în domeniul managementului sănătăţii publice, precum şi componentele managementului instituţional al Şcolii de Management în Sănătate Publică, în calitate de promotor al noilor abordări în domeniu; rolul, valoarea şi perspectivele de dezvoltare ale instituţiei.Beginning with 2001, The Ministry of Health, local and international decision-makers initiated the achievement of the capacity building objectives in Public Health Management, by creating the School of Public Health Management in the Republic of Moldova, that started its activity in 2003. The present paper approaches the total capacities development efforts in the area of public health over the last decade (2005-2015), in particular by organizing the Masters program in the nominated field in the Republic of Moldova. The elucidated aspects include training components, research, services in public health management, as well as institutional management components of the School of Public Health Management, as a promoter of new approaches in the field; the role, value and development prospects of the institution

The LDB1 Complex Co-opts CTCF for Erythroid Lineage-Specific Long-Range Enhancer Interactions

Summary: Lineage-specific transcription factors are critical for long-range enhancer interactions, but direct or indirect contributions of architectural proteins such as CCCTC-binding factor (CTCF) to enhancer function remain less clear. The LDB1 complex mediates enhancer-gene interactions at the β-globin locus through LDB1 self-interaction. We find that an LDB1-bound enhancer upstream of carbonic anhydrase 2 (Car2) activates its expression by interacting directly with CTCF at the gene promoter. Both LDB1 and CTCF are required for enhancer-Car2 looping, and the domain of LDB1 contacted by CTCF is necessary to rescue Car2 transcription in LDB1-deficient cells. Genome-wide studies and CRISPR/Cas9 genome editing indicate that LDB1-CTCF enhancer looping underlies activation of a substantial fraction of erythroid genes. Our results provide a mechanism by which long-range interactions of architectural protein CTCF can be tailored to achieve a tissue-restricted pattern of chromatin loops and gene expression. : It is unclear whether or how architectural proteins such as CTCF might contribute to interactions between enhancers and target genes. Lee et al. show that CTCF interacts directly with LDB1 and that a significant fraction of erythroid genes loop to LDB1-complex-occupied enhancers via CTCF sites within their promoters. Keywords: LDB1, CTCF, enhancer, looping, erythroid cell