Alemany Group - Stochastic Choice and Regulation

Epigenetic effects in 2D directed differentiation protocols

Induced pluripotent stem cells (iPSCs) were derived from human fibroblasts for the first time in 2007, triggering our collective imagination in the field of personalized regenerative medicine. Nowadays, iPSCs can also be derived from other somatic cells and differentiated to a broad range of cell types present in adult tissues. However, iPSCs are still not being widely used in personalized therapies. Why is that?

iPSCs have several sources of variability. For example, the DNA genetic code of iPSCs varies from line to line because of the patient-specific genetic mutations. In addition, epigenetic effects related to the somatic origin used to derive each line can bias their differentiation potential.

In our lab, we are combining RNA, ATAC and methylation sequencing to quantitatively characterize the role of variability in iPSC differentiation potential towards different lineages. We work in collaboration with the IPS and LGTC facilites at the LUMC, and Diagenode (Belgium), with the aim to reconstruct the differentiation landscape of distinct patient-derived iPSC lines and hope to find common predictive genetic and epigenetic features.

Quantitative characterization of gene regulatory networks during cell fate commitment

During differentiation, cells usually undergo bifurcation events where they “decide” between two or more fates. A widespread situation has been observed, in which bi-potent progenitors co-express two genes (or transcription factors, TF), with each being repressed as commitment to one or another fate takes place.

Here, we propose scenario in which gene expression patterns are regulated by gene regulatory networks (GRN): the fate of a bi-potent progenitor follows probabilistic rules set by the GRNs. To test our hypotheses, we focus on the characterization of the regulatory network underlying neuro-mesodermal progenitor (NMP) commitment towards the mesodermal or neuronal fates in in vivo and in vitro systems. We are adapting current inducible lineage molecular recorders to uniquely label cells with different degrees of TF activity during fate commitment. The recorder read-out is combined with transcriptome detection with single-cell resolution and mathematical modeling, allowing us to quantify the role gene-gene interactions during cell decision making.

Reconstruction of signaling histories in single cells

Cells differentiate as a result of the integration between extracellular signals and gene expression programs. A better understanding of the sequence and intensities of signaling events that cells experience during fate commitment (in embryonic development, but also disease context) will lead to more effective designs of in vitro differentiation protocols and drug therapies.

By using inducible CRISPR/Cas9-based systems, we plan to record the signaling histories of single cells undergoing targeted differentiation. From our results plan to elucidate the kinetic properties of signaling pathway activation and design new differentiation protocols effectively.