Perturbing haematopoietic transcription factor networks and cell fate decisions using transcriptional activator-like effectors

Transcription factors are key determinants of cell identity and fate. Numerous TFs are known to play critical roles in haematopoiesis, including Scl/Tal1, PU.1 and Gata1. Our laboratory, along with others, has identified many transcriptional cis-regulatory elements within gene loci of key haematopoietic TFs, often distal to gene promoters. Haematopoietic TFs interact via these regulatory elements to form TF circuits and interconnected networks that act to antagonise or reinforce cell fate decisions. Transcriptional Activator-Like Effector (TALE) proteins have modular and predictable DNA binding domains, a feature that allows their de novo assembly as synthetic sequence-specific TFs. Fusion of transcriptional activator (VP64) or repressor (KRAB) domains to TALEs targeting promoters has recently been shown to modulate gene expression. We have designed TALE activators and repressors to target several haematopoietic TF regulatory elements, including the Scl+40kb, PU.1-14kb and Gata1-3.7kb enhancers. Initial testing in haematopoietic cell lines confirmed their ability to modulate target gene expression from these distal regulatory elements. Interestingly, expression of neighbouring genes can also affected suggesting their co-regulation. We have further applied TALEs to haematopoietic programming from embryonic stem cells and analysed their perturbation of TF networks in haematopoietic progenitor cell fate decisions. We will present validation of TALE activators and repressors targeting distal regulatory elements as an efficient method of perturbing haematopoietic TF networks and the novel insights into cell fate decisions gained.