A small-molecule screen reveals that HSP90beta promotes the conversion of induced pluripotent stem cell-derived endoderm to a hepatic fate and regulates HNF4A turnover

We have previously shown that the transcription factor HNF4A is required for the formation of hepatic progenitor cells from endoderm that has been derived from human induced pluripotent stem cells (iPSCs). We reasoned that we could uncover regulatory pathways with new roles in hepatocyte differentiation by identifying cellular processes that regulate HNF4A. We therefore performed a screen of 1120 small molecules with well-characterized mechanisms of action to detect those that affect the abundance of HNF4A in iPSC-derived hepatic progenitor cells. This approach uncovered several small molecules that depleted HNF4A. Of those, we chose to focus on an inhibitor of heat shock protein 90 beta (HSP90beta). We show that mutation of the gene encoding HSP90beta represses hepatocyte differentiation during the formation of hepatocytes from iPSCs. We reveal that HSP90beta, although dispensable for expression of HNF4A mRNA, directly interacts with HNF4A protein to regulate its half-life. Our results demonstrate that HSP90beta has an unappreciated role in controlling hepatic progenitor cell formation and highlight the efficiency of using small-molecule screens during the differentiation of iPSCs to reveal new molecular mechanisms that control hepatocyte formation.