NovoSpaRc: flexible spatial reconstruction of single-cell gene expression with optimal transport.

Single-cell RNA-sequencing (scRNA-seq) technologies have revolutionized modern biomedical sciences. A fundamental challenge is to incorporate spatial information to study tissue organization and spatial gene expression patterns. Here, we describe a detailed protocol for using novoSpaRc, a computational framework that probabilistically assigns cells to tissue locations. At the core of this framework lies a structural correspondence hypothesis, that cells in physical proximity share similar gene expression profiles. Given scRNA-seq data, novoSpaRc spatially reconstructs tissues based on this hypothesis, and optionally, by including a reference atlas of marker genes to improve reconstruction. We describe the novoSpaRc algorithm, and its implementation in an open-source Python package ( https://pypi.org/project/novosparc ). NovoSpaRc maps a scRNA-seq dataset of 10,000 cells onto 1,000 locations in <5 min. We describe results obtained using novoSpaRc to reconstruct the mouse organ of Corti de novo based on the structural correspondence assumption and human osteosarcoma cultured cells based on marker gene information, and provide a step-by-step guide to Drosophila embryo reconstruction in the Procedure to demonstrate how these two strategies can be combined.

Human muscle-derived CLEC14A-positive cells regenerate muscle independent of PAX7.

Skeletal muscle stem cells, called satellite cells and defined by the transcription factor PAX7, are responsible for postnatal muscle growth, homeostasis and regeneration. Attempts to utilize the regenerative potential of muscle stem cells for therapeutic purposes so far failed. We previously established the existence of human PAX7-positive cell colonies with high regenerative potential. We now identified PAX7-negative human muscle-derived cell colonies also positive for the myogenic markers desmin and MYF5. These include cells from a patient with a homozygous PAX7 c.86-1G > A mutation (PAX7null). Single cell and bulk transcriptome analysis show high intra- and inter-donor heterogeneity and reveal the endothelial cell marker CLEC14A to be highly expressed in PAX7null cells. All PAX7-negative cell populations, including PAX7null, form myofibers after transplantation into mice, and regenerate muscle after reinjury. Transplanted PAX7neg cells repopulate the satellite cell niche where they re-express PAX7, or, strikingly, CLEC14A. In conclusion, transplanted human cells do not depend on PAX7 for muscle regeneration.