Uncovering the transcription factor networks in early human cell specification. During gastrulation, pluripotent epiblast cells give rise to the three germ layers-- endoderm, mesoderm, and ectoderm. Later in development, these layers form nearly all the different tissues and organs in our body. However the molecular mechanisms that establish the transition from epiblast to the three germ layers are still largely unknown. A class of proteins known as transcription factors (TFs) can bind specific DNA elements and activate or repress gene expression. The capacity of TFs to "de-differentiate" fibroblast cells to induced pluripotent stem cells (1, 2) or to directly program cells into various lineages (3) makes them likely candidates for regulating cellular transitions during development. To gain a deeper understanding of the regulatory events that guide early human cell specification, a more comprehensive study of TF binding and their dynamics is needed. Current research on TFs in development mainly focuses on the regulatory role of single factors in steady state conditions, yet many TFs mediate gene expression downstream of signaling cascades in a dynamic and cooperative fashion that is unique to each cell type (4, 5). During my postdoctoral studies I aim to uncover the molecular events underling cellular specification. First, I will determine the genome-wide dynamics of over 30 TFs, DNA methylation, chromatin marks, and RNA expression at multiple decision time-points, during differentiation of human embryonic stem (ES) cells into endoderm, mesoderm, and ectoderm. Such comprehensive maps of TF binding dynamics will allow me to dissect the combinatorial and temporal interactions between master regulators, cofactors, and signaling proteins that establish cell identity. Second, I will combine these dynamic measurements to generate a provisional model of the network that controls cell identity in the different germ layers and dissect the interplay between TF occupancy and epigenetic state in the regulation of development. Third, I will validate and reiterate my multi-dimensional model predictions using selected RNAi perturbations of critical TFs in the network. The combination of this data will allow me to uncover the principles and players that establish cell fate. The proposed research will greatly enhance our understanding of regulatory circuits and their roles during cell differentiation in early human development, which will, in turn, improv our ability to derive therapeutic approaches, such as in vitro generation of cardiomyocytes, pancreatic islets, and neurons. This promises to have significant impact in combating number of diseases, such as spinal cord injury, juvenile diabetes, and Parkinson's disease.
As a postdoctoral fellow, I plan to dissect the transcription factor (TF) networks that establish cell fate in early human development. I will combine TF binding, RNA expression, and DNA methylation data to model how combinatorial relationships between dozens of TFs guide the differentiation of cells;using siRNA perturbation I will validate, reiterate and improve on the model. The proposed study will greatly improve our understanding of lineage specification and will provide a critical step forward in engineering in vitro cell type and in regenerative medicine.
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|Ziller, Michael J; Edri, Reuven; Yaffe, Yakey et al. (2015) Dissecting neural differentiation regulatory networks through epigenetic footprinting. Nature 518:355-9|
|Tsankov, Alexander M; Akopian, Veronika; Pop, Ramona et al. (2015) A qPCR ScoreCard quantifies the differentiation potential of human pluripotent stem cells. Nat Biotechnol 33:1182-92|
|Tsankov, Alexander M; Gu, Hongcang; Akopian, Veronika et al. (2015) Transcription factor binding dynamics during human ES cell differentiation. Nature 518:344-9|
|Gifford, Casey A; Ziller, Michael J; Gu, Hongcang et al. (2013) Transcriptional and epigenetic dynamics during specification of human embryonic stem cells. Cell 153:1149-63|