ABSTRACT: The lipodystrophic hotspot lamin A p.R482W mutation deregulates the mesodermal inducer T/Brachyury and early vascular differentiation gene networks
Project description:The p.R482W hotspot mutation in A-type nuclear lamins causes familial partial lipodystrophy of Dunnigantype (FPLD2), a lipodystrophic syndrome complicated by early-onset atherosclerosis. Molecular mechanisms underlying endothelial cell dysfunction conferred by the lamin A mutation remain elusive. However, lamin A regulates epigenetic developmental pathways and mutations could perturb these functions. Here, we demonstrate that lamin A R482W elicits endothelial differentiation defects in a developmental model of FPLD2. Genome modeling in fibroblasts from patients with FPLD2 caused by the lamin A R482W mutation reveals repositioning of the mesodermal regulator T/Brachyury locus towards the nuclear center relative to normal fibroblasts, suggesting enhanced activation propensity of the locus in a developmental model of FPLD2. Addressing this issue, we report phenotypic and transcriptional alterations in mesodermal and endothelial differentiation of induced pluripotent stem cells we generated from a patient with R482Wassociated FPLD2. Correction of the LMNA mutation ameliorates R482W-associated phenotypes and gene expression. Transcriptomics links endothelial differentiation defects to decreased Polycomb-mediated repression of the T/Brachyury locus and over-activation of T target genes. Binding of the Polycomb repressor complex PRC2 to T/Brachyury is impaired by the mutated lamin A network, which is unable to properly associate with the locus. This leads to a deregulation of vascular gene expression over time. By connecting a lipodystrophic hotspot lamin A mutation to a disruption of early mesodermal gene expression and defective endothelial differentiation, we propose that the mutation rewires the fate of several lineages, resulting in multitissue pathogenic phenotypes.
Project description:Brachyury (Xbra/Xbra3) knock-down embryos of the frog Xenopus tropicalis were profiled to quantify neuro-mesodermal cell fate switches at a transcriptional level. Transcriptional profiling of Xbra/Xbra3 double morphants at early tadpole stage (RNA-Seq) in biological triplicates.
Project description:ChIP-seq to clarify the molecular mechanism by which a mesodermal transcription factor, T/Brachyury, disrupts the defined naive pluripotency.
Project description:In vertebrate embryos, anterior tissues are generated early, followed by the other axial structures that emerge sequentially from a posterior growth zone. The genetic network driving posterior axial elongation in mice, and its disturbance in mutants with posterior truncation are not yet fully understood. We show that the combined expression of Cdx2 and T Brachyury is essential to establish the core signature of posterior axial progenitors. Cdx2 and T Brachyury are required for extension of a similar trunk portion of the axis. Simultaneous loss of function of these two genes disrupts axial elongation to a much greater extent than each single mutation alone. We identify and validate common targets for Cdx2 and T Brachyury in vivo including Wnt and Fgf pathway components active in the axial progenitor niche. Our data demonstrate that integration of the Cdx/Hox and T Brachyury transcriptional networks controls differential axial growth during vertebrate trunk elongation.
Project description:β-Catenin is the major co-regulator of the Wnt signalling pathway. β-Catenin lysine 49 is post-translationally modified. The histone methyl transferase Ezh2 trimethylates β-catenin at lysine 49 and the acetyl transferase Cbp acetylates β-catenin at the same lysine. To determine the effects on gene expression embryonic stem cells containing either a Gfp tagged β-catenin wildtype (wt) or lysine 49 to alanine (K49A) loss of function mutation were analysed by micro array expression profiling. To determine the effects on gene expression in the pluripotent state two biological replicates of Gfp β-catenin wt and Gfp β-catenin K49A were analysed. The results showed that genes which affect pluripotency as well as differentiation were altered by β-catenin K49A mutation. To analyse the effects of gene expression during ES cell differentiation ES cells containing Gfp β-catenin wt or K49A were differentiated into mesodermal progenitors (mp) and neuronal progenitors (np). Two biological replicates were analysed by micro array expression profiling. Differentially expressed genes between Gfp β-catenin wt and K49A during differentiation were observed. Mesodermal marker genes like t-brachyury and cdx2 were not upregulated in mesodermal differentiation of Gfp β-catenin K49A Es cells.
Project description:RNA-seq to clarify the molecular mechanism by which a mesodermal transcription factor, T/Brachyury, disrupts the defined naive pluripotency and induce the concomitant differentiation program.
Project description:Brachyury (Xbra/Xbra3) knock-down embryos of the frog Xenopus tropicalis were profiled to quantify neuro-mesodermal cell fate switches at a transcriptional level.
Project description:We defined genome-wide regulatory inputs of the T-box transcription factors Brachyury (Xbra), Eomesodermin (Eomes) and VegT that maintain neuro-mesodermal stem cells and determine their bipotential fates in the Xenopus tropicalis frog embryo.
Project description:We identified distict mesodermal sub-populations based on Endoglin (Eng) and Flk1 expression in Brachyury (Bry) positive cells. By using whole-transcriptome analysis, we further characterized these populations and how they changed when Wnt pathway is inhibited
Project description:We defined genome-wide regulatory inputs of the T-box transcription factors Brachyury (Xbra), Eomesodermin (Eomes) and VegT that maintain neuro-mesodermal stem cells and determine their bipotential fates in the Xenopus tropicalis frog embryo. Binding profiles for Xbra, Eomes and VegT in X. tropicalis embryos (ChIP-Seq)
