Single-Cell Sequencing and in Vivo Analysis of the Biliary Epithelium Reveal a Dynamic and Essential Role for Yap

Abstract

The liver, which serves as the main filtration system for the body, is composed of a variety of cell types. Biliary epithelial cells (BECs), an essential cell type within the liver, serve as the main structural component to bile ducts and are fundamental to metabolism through their regulation of the composition and flow of bile. In addition to facilitating metabolic processes, they serve as a principal cell source for liver regeneration after injury, even acting as facultative liver progenitor cells to repopulate the liver parenchyma in situations of prolonged injury. Although BECs have been suggested to be functionally and transcriptionally heterogeneous, an impartial analysis of the genetic diversity within this population has yet to be explored. Here, we harness the power of single-cell sequencing to analyze adult BECs under homeostatic conditions, thereby unearthing the critical regulatory pathways that define this essential population. In doing so, we identify a unique subset of adult BECs that express genes associated with the activated form of the co- transcriptional regulator Yap, a primary controller of cell growth, stem-ness, and ductular expansion in the liver. Furthermore, we find that this gene signature is dynamic and can be activated upon environmental stimuli and injury. After identifying this signature, we probe the role of Yap in the regulation of adult BECs, and demonstrate for the first time, using two genetic mouse models, that Yap is essential for BEC maintenance. Indeed, inducible loss of Yap in vivo, leads to the dramatic loss of BECs, an increase of cell death genes, and a decrease in the originally identified Yap associated gene signature. Finally, we demonstrate that this Yap-induced signature is driven, in part, in response to the normal inflow of toxic bile acids and is attenuated upon administration of a bile acid sequestrant, resin. Thus, our pioneering work provides a robust framework for understanding signalling heterogeneity within cells of the normal and injured biliary tree, reveals a unique requirement for Yap within the adult biliary epithelium, and exposes a role for Yap in BECs as a protective rheostat in dynamically responding to environmental stimuli.