Project description:We report the application of Chromosome Conformation Capture Carbon-copy (5C) to a 4.5 Mb stretch of the mouse X chromosome encompassing the X inactivation center locus. We uncover a series of discrete 200kb-1Mb topologically associating domains (TADs). These align with several domain-wide epigenomic features as well as co-regulated gene clusters. 5C analysis in EED and G9A mutants reveal that this segmental organisation in TADs does not relie on the underlying H3K27me3 or H3K9me2 blocks. Deletion of a boundary between two TADs leads to ectopic chromosomal contacts between them. Analysis of mESCs, mNPCs and MEFs suggest that the positioning of TADs on the chromosome is stable during cell differentiation though their internal organisation changes. Comparison of male (XY) and female (XX) differentiated cells highlights that the long-range chromosomal contacts within TADs are dampened on the inactive X compared to the active X. 5C oligonucleotides were designed around HindIII restriction site following an alternative scheme
Project description:Heart failure affects 2–3% of adult Western population. Prevalence of heart failure with preserved left ventricular (LV) ejection fraction (HFpEF) increases. Studies suggest HFpEF patients to have altered myocardial structure and functional changes such as incomplete relaxation and increased cardiac stiffness. We hypothesised that patients undergoing elective coronary bypass surgery (CABG) with HFpEF characteristics will show distinctive gene expression compared to patients with normal LV physiology. Myocardial biopsies for mRNA expression analysis were obtained from sixteen patients with LV ejection fraction ≥ 45%. Five out of 16 patients (31%) had echocardiographic characteristics and increased NTproBNP levels indicative of HFpEF and this group was used as HFpEF proxy, while 11 patients had Normal LV physiology. Utilising principal component analysis, the gene expression data clustered into two groups, corresponding to HFpEF proxy and Normal physiology, and 743 differentially expressed genes were identified. The associated top biological functions were cardiac muscle contraction, oxidative phosphorylation, cellular remodelling and matrix organisation. Our results also indicate that upstream regulatory events, including inhibition of transcription factors STAT4, SRF and TP53, and activation of transcription repressors HEY2 and KDM5A, could provide explanatory mechanisms to observed gene expression differences and ultimately cardiac dysfunction in the HFpEF proxy group. <br> Sequencing data from clinical patients fall under GDPR regulations of sharing of personal data and will be made available through EGA-SE.
Project description:We report the application of Chromosome Conformation Capture Carbon-copy (5C) to a 4.5 Mb stretch of the mouse X chromosome encompassing the X inactivation center locus. We uncover a series of discrete 200kb-1Mb topologically associating domains (TADs). These align with several domain-wide epigenomic features as well as co-regulated gene clusters. 5C analysis in EED and G9A mutants reveal that this segmental organisation in TADs does not relie on the underlying H3K27me3 or H3K9me2 blocks. Deletion of a boundary between two TADs leads to ectopic chromosomal contacts between them. Analysis of mESCs, mNPCs and MEFs suggest that the positioning of TADs on the chromosome is stable during cell differentiation though their internal organisation changes. Comparison of male (XY) and female (XX) differentiated cells highlights that the long-range chromosomal contacts within TADs are dampened on the inactive X compared to the active X.