Project description:We have determined the genome-wide binding profile of the transcription factor Sp5. Flag-tagged Sp5 was targeted to the HPRT locus in A2Lox.cre ES cells to allow for Doxycycline inducible expression. ES cells were cultured as embryoid bodies for 2 days to prime them for germ layer differentiation. Cells were then treated with Doxycycline for 24 hrs. We found that Flag-Sp5 prefentially bound to promoters associatd with several growth factor signalling pathways, but most prominently with the Wnt/beta-catenin pathway to selectively promote mesoderm differentiation over neural. These data implicate Sp5 as new regulator of Wnt/beta-catenin target expression to promotes ES cell differentation into the mesoderm lineage. Examination of Sp5 transcription factor binding in differentiating embryonic stem cells.
Project description:During in vitro differentiation, pluripotent stem cells undergo extensive remodeling of their gene expression profiles. While studied extensively at the transcriptome level, much less is known about protein dynamics, which might differ significantly from their mRNA counterparts. Here, we present deep proteome-wide measurements of protein levels during the differentiation of embryonic stem cells.
Project description:The Nucleosome Remodeling and Deacetylase (NuRD) complex plays an important role in gene expression regulation, stem cell self-renewal, and lineage commitment. Yet little is known about the dynamics of NuRD during cellular differentiation. Here, we study these dynamics using genome-wide profiling and quantitative interaction proteomics in mouse embryonic stem cells (ESCs) and neural progenitor cells (NPCs). The genomic targets of NuRD are highly dynamic during differentiation, with most binding occurring at cell-type specific promoters and enhancers. We identify ZFP296 as a novel, ESC-specific NuRD interactor that also interacts with the SIN3A complex. ChIP-sequencing in Zfp296 knockout (KO) ESCs reveals decreased NuRD binding both genome-wide and at ZFP296 binding sites, although this has little effect on the transcriptome. Nevertheless, Zfp296 KO ESCs exhibit delayed induction of lineage-specific markers upon differentiation to embryoid bodies. In summary, we identify an ESC-specific NuRD interacting protein which regulates genome-wide NuRD binding and cellular differentiation.
Project description:Genome-wide binding profiles of nuclear FGFR1, RXRα and Nur77 in pluripotent mouse Embryonic Stem Cells and during Retinoic Acid-induced differentiation
Project description:The WNT/β-catenin signaling pathway is a prominent player in many developmental processes, including gastrulation, anterior-posterior axis specification, organ and tissue development and homeostasis. Here, we use human pluripotent stem cells (hPSCs) to study the dynamics of the transcriptional response to exogenous activation of the WNT pathway. We describe a mechanism involving the WNT target gene SP5 that leads to termination of the transcriptional program initiated by WNT signaling. Integration of gene expression profiles of wildtype and SP5 mutant cells with genome-wide SP5 binding events reveals that SP5 acts to diminish expression of genes previously activated by the WNT pathway. Furthermore, we show that activation of SP5 by WNT signaling is most robust in cells with developmental potential, such as stem cells. These findings indicate a mechanism by which the developmental WNT signaling pathway reins in expression of transcriptional programs.