Executive Director Allen Institute for Cell Science Seattle, Washington, United States
Abstract: The Allen Institute for Cell Science aims to understand the principles by which human induced pluripotent stem cells (hiPSCs) establish and maintain robust dynamic localization of cellular structures, and how they transition between states during differentiation and disease. To accomplish these goals, we used CRISPR/Cas9 to create a collection of endogenously tagged hiPSC lines to illuminate organization in various cell states from pluripotency through differentiation. The tagged proteins include notable membrane-bound and membrane-less cellular organelles, signaling complexes, phase transition markers, transcription factors and differentiation-specific structural and cell identity markers. To date, the Allen Cell Collection consists of 56 tagged cell lines that have undergone extensive quality control testing to ensure genomic, cell biological, and stem cell integrity. Here, we describe the development and application of these cell lines with live dynamic 3D imaging to study key biological processes including cell division, migration, and differentiation into distinct cell types. For example, we have generated TBR2-mEGFP (EOMES) and TBXT-Halo (Brachyury) to follow the early differentiation process during the epithelial-mesenchymal transition, CDH5-mEGFP (Vascular E-cadherin) to characterize endothelial cells, and MYH7-mEGFP (Myosin heavy chain 7) to visualize myofibrils in cardiomyocytes. These endogenously tagged hiPSC lines have allowed us to study the intracellular organization, heterogeneity, and dynamics of various cell states and cell state transitions over time. In addition, we have generated 8 disease cell line collections to study cardiac, skeletal and nuclear disease phenotypes. We will describe the workflows used to generate these collections and highlight some of the applications leading to novel insights on disease progression. Our cell lines, plasmids, 3D images, image analysis and visualization tools, integrated cell models and biological findings are openly available for the research community (www.allencell.org).
