Transcription factor-based reprogramming of differentiated cells to induced pluripotent stem cells (iPSCs) has revolutionized the field of stem cell biology and greatly increased the opportunities for disease modeling and cell-based regenerative therapies. While iPSCs offer tremendous potential, a number of technical challenges remain in the generation of iPSCs that hamper their use in clinical settings. A comprehensive understanding of the molecular mechanisms of cellular reprogramming is essential for devising methods to overcome these challenges. One of the fundamental questions with respect to these mechanisms is how somatic cell identity gets erased and then replaced with a self-sustaining pluripotency network. Maintenance of cellular identity is controlled by epigenetic mechanisms that are based on chromatin marks including DNA methylation and histone modifications. Remodelling the somatic cell chromatin structures poses a major barrier for the activation of pluripotency-specific gene expression during reprogramming. Our research is aimed at understanding the molecular mechanisms of reprogramming and identifying genes that are important for iPSC generation. Specifically, we utilize forward genetics tools (i.e. shRNA and CRISPR-Cas9 technology) and small molecule inhibitors to identify the role chromatin modifying enzymes play during reprogramming.
Our long term research goal is to understand the mechanisms that maintain cell identity and determine whether perturbations of these pathways can result in efficient and faithful reprograming to pluripotency.
Understanding the Role of Chromatin Modifiers in Somatic Cell Reprogramming
In recent work, we have employed a loss-of-function approach to interrogate the role of genes in DNA and histone methylation pathways and identified Dot1L, the only known H3K79 methyl-transferase, as a potent suppressor of cellular reprogramming. Inhibition of Dot1L either by RNA interference or a small molecule inhibitor accelerated reprogramming, significantly increased the yield of iPSCs, and substituted for two of the exogenous transcription factors used in reprogramming . To determine the role of additional chromatin modifiers in reprogramming we are currently pursuing a combination of candidate-based and high-throughput molecular approaches.