Professor Keisuke Kaji, MRC Centre for Regenerative Medicine, University of Edinburgh

Title: Molecular mechanisms of cellular reprogramming

Abstract:

The generation of induced pluripotent stem cells (iPSCs) by overexpression of Oct4, Sox2, Klf4 and c-Myc, transformed our classical views of the cellular epigenetic landscape and highlighted transcription factors as powerful tools for cell conversions and tissue engineering. Following this seminal work, several other cell types have also been generated by master transcription factor (TF)-mediated transdifferentiation. However, the molecular mechanisms underlying the diverse cellular identity changes are not well understood. Through the investigation of reprogramming mechanisms, we recently revealed that overexpression of constitutive active Smad3 enhanced not only iPSC generation, but also 3 other master TF-mediated conversions, from B cells to macrophages, myoblasts to adipocytes, and human fibroblasts to neurons. This has demonstrated that there are common mechanisms underlying different master TF-mediated cell conversions. To illuminate reprogramming mechanisms further, we have performed CRISPR/Cas9-mediated genome-wide knockout screening during MEF reprogramming with a lentiviral gRNA library containing 90,000 gRNAs. This screening provided us with 16 novel reprogramming roadblock genes as well as 16 genes essential for iPSC generation but not for ES cell self-renewal or MEF proliferation. Excitingly, many of those essential genes enhanced reprogramming when overexpressed together with Yamanaka factors. This data set will be a valuable resource for better understanding of molecular mechanisms of iPSC generation, and which could be influential in achieving any other cell conversions more efficiently and faithfully.