Abstract

Chemical Biology and High-throughput Strategy for Dissecting Human Stem Cell Pluripotency and Differentiation for Translational Research

Jiayu Liao, Associate Professor, University of California Riverside

Understanding and controlling the fate decisions of human pluripotent stem cells (hPSCs) is central to human developmental studies and the potential applications for disease modeling, drug discovery and cell-based therapies. We have developed a general high-throughput screening and follow-up chemical biology strategy to dissecting the complex network that control human stem cell pluripotency and its relationships to differentiations into different cell lineages. This strategy is mainly based on our fluorescent protein reporter genes driven by the promoter of Oct4 that is essential for human stem cell pluripotency.  We demonstrate an essential role for the serine/threonine protein kinase mammalian target of rapamycin (mTOR) in maintaining hESC undifferentiated growth. mTOR integrates signals from extrinsic pluripotency-supporting factors and represses the transcriptional activity of a subset of developmental and growth-inhibitory genes. Furthermore, transcriptional repression of the developmental genes by mTOR is independent of Oct-4, Sox-2 and Nanog and is mediated partially by suppression of the Wnt signaling pathway. In a recent large scale screening against >170,000 compounds, leading to the identification of more than a hundred of lead chemicals that potently disrupt cellular pluripotency. By exploiting target identification, biochemical, molecular and cellular analyses, we discover a novel small-molecule inhibitor that promotes transcription circuitry of pluripotency and another lead chemical, when used in conjunction with extrinsic factors, leads to robust, close-to-homogeneity mesoderm and endoderm differentiation of human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs). Together, we provide novel mechanistic insights into pluripotency and establish new tools and technologies for monitoring and manipulating hPSC fate, which should greatly facilitate the biological and therapeutic applications of hPSCs.