Tracing the lineage history of cells is key to answering diverse and fundamental questions in biology. Particularly in the context of stem cell biology, analysis of single cell lineages in their native state has elucidated novel fates and highlighted heterogeneity of function. Coupling of such ancestry information with other molecular readouts represents an important goal in the field. Here, we describe the CARLIN (for CRISPR Array Repair LINeage tracing) mouse line and corresponding analysis tools that can be used to simultaneously interrogate the lineage and transcriptomic information of single cells in vivo. This model exploits CRISPR technology to generate up to 44,000 transcribed barcodes in an inducible fashion at any point during development or adulthood, is compatible with sequential barcoding, and is fully genetically defined. We have used CARLIN to identify intrinsic biases in the activity of fetal liver hematopoietic stem cell (HSC) clones and to uncover a previously unappreciated clonal bottleneck in the response of HSCs to injury. CARLIN also allows the unbiased identification of transcriptional signatures based on in vivo stem cell function without a need for markers or cell sorting..

Highlights d Hippo signaling inhibition reprograms Lgr5+ ISCs to a Klf6+ wound-healing cell state d Klf6+ cells are characterized by low Wnt activity and reduced stemness d YAP activation suppresses tumor growth in human and mouse metastatic CRC d YAP/TAZ deletion increases colon tumor growth

SummarySkeletal muscle harbors quiescent stem cells termed satellite cells and proliferative progenitors termed myoblasts, which play pivotal roles during muscle regeneration. However, current technology does not allow permanent capture of these cell populations in vitro. Here, we show that ectopic expression of the myogenic transcription factor MyoD, combined with exposure to small molecules, reprograms mouse fibroblasts into expandable induced myogenic progenitor cells (iMPCs). iMPCs express key skeletal muscle stem and progenitor cell markers including Pax7 and Myf5 and give rise to dystrophin-expressing myofibers upon transplantation in vivo. Notably, a subset of transplanted iMPCs maintain Pax7 expression and sustain serial regenerative responses. Similar to satellite cells, iMPCs originate from Pax7+ cells and require Pax7 itself for maintenance. Finally, we show that myogenic progenitor cell lines can be established from muscle tissue following small-molecule exposure alone. This study thus reports on a robust approach to derive expandable myogenic stem/progenitor-like cells from multiple cell types.