Functionally heterogeneous human satellite cells identified by single cell RNA sequencing
- University of California, San Francisco, United States
Abstract
Although heterogeneity is recognized within the murine satellite cell pool, a comprehensive understanding of distinct subpopulations and their functional relevance in human satellite cells is lacking. We used a combination of single cell RNA sequencing and flow cytometry to identify, distinguish, and physically separate novel subpopulations of human PAX7+ satellite cells (Hu-MuSCs) from normal muscles. We found that, although relatively homogeneous compared to activated satellite cells and committed progenitors, the Hu-MuSC pool contains clusters of transcriptionally distinct cells with consistency across human individuals. New surface marker combinations were enriched in transcriptional subclusters, including a subpopulation of Hu-MuSCs marked by CXCR4/CD29/CD56/CAV1 (CAV1+). In vitro, CAV1+ Hu-MuSCs are morphologically distinct, and characterized by resistance to activation compared to CAV1- Hu-MuSCs. In vivo, CAV1+ Hu-MuSCs demonstrated increased engraftment after transplantation. Our findings provide a comprehensive transcriptional view of normal Hu-MuSCs and describe new heterogeneity, enabling separation of functionally distinct human satellite cell subpopulations.
Data availability
Single cell rna sequencing data were uploaded to Dryad and can be accessed under the doi: 10.7272/Q65X273X
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Functionally heterogeneous human satellite cells identified by single cell RNA sequencingDryad Digital Repository, 10.7272/Q65X273X.
Article and author information
Author details
Funding
California Institute for Regenerative Medicine (New Faculty Physician Scientist Award RN3-06504)
- Jason H Pomerantz
National Institutes of Health (R01AR072638-03)
- Jason H Pomerantz
University of California, San Francisco (UCSF PROF-PATH program via NIH R25MD006832)
- Steven M Garcia
University of California, San Francisco (Research Allocation Program for trainees)
- Solomon Lee
Eli and Edythe Broad Foundation (Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research Fellowship)
- Alvin Wong
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Shahragim Tajbakhsh, Institut Pasteur, France
Ethics
Animal experimentation: All procedures were approved and performed in accordance with the UCSF Institutional Animal Care and Use Committee (Protocols #181101).
Human subjects: This study was conducted under the approval of the Institutional Review Board at The University of California San Francisco (UCSF). Written informed consent was obtained from all subjects.
Version history
- Received: September 4, 2019
- Accepted: March 27, 2020
- Accepted Manuscript published: April 1, 2020 (version 1)
- Version of Record published: April 17, 2020 (version 2)
Copyright
© 2020, Barruet et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Functionally heterogeneous human satellite cells identified by single cell RNA sequencing
eLife 9:e51576.
https://doi.org/10.7554/eLife.51576
Further reading
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- Developmental Biology
- Stem Cells and Regenerative Medicine
Endothelial and skeletal muscle lineages arise from common embryonic progenitors. Despite their shared developmental origin, adult endothelial cells (ECs) and muscle stem cells (MuSCs; satellite cells) have been thought to possess distinct gene signatures and signaling pathways. Here, we shift this paradigm by uncovering how adult MuSC behavior is affected by the expression of a subset of EC transcripts. We used several computational analyses including single-cell RNA-seq (scRNA-seq) to show that MuSCs express low levels of canonical EC markers in mice. We demonstrate that MuSC survival is regulated by one such prototypic endothelial signaling pathway (VEGFA-FLT1). Using pharmacological and genetic gain- and loss-of-function studies, we identify the FLT1-AKT1 axis as the key effector underlying VEGFA-mediated regulation of MuSC survival. All together, our data support that the VEGFA-FLT1-AKT1 pathway promotes MuSC survival during muscle regeneration, and highlights how the minor expression of select transcripts is sufficient for affecting cell behavior.
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- Biochemistry and Chemical Biology
- Stem Cells and Regenerative Medicine
Molecules that facilitate targeted protein degradation (TPD) offer great promise as novel therapeutics. The human hepatic lectin asialoglycoprotein receptor (ASGR) is selectively expressed on hepatocytes. We have previously engineered an anti-ASGR1 antibody-mutant RSPO2 (RSPO2RA) fusion protein (called SWEETS) to drive tissue-specific degradation of ZNRF3/RNF43 E3 ubiquitin ligases, which achieved hepatocyte-specific enhanced Wnt signaling, proliferation, and restored liver function in mouse models, and an antibody–RSPO2RA fusion molecule is currently in human clinical trials. In the current study, we identified two new ASGR1- and ASGR1/2-specific antibodies, 8M24 and 8G8. High-resolution crystal structures of ASGR1:8M24 and ASGR2:8G8 complexes revealed that these antibodies bind to distinct epitopes on opposing sides of ASGR, away from the substrate-binding site. Both antibodies enhanced Wnt activity when assembled as SWEETS molecules with RSPO2RA through specific effects sequestering E3 ligases. In addition, 8M24-RSPO2RA and 8G8-RSPO2RA efficiently downregulate ASGR1 through TPD mechanisms. These results demonstrate the possibility of combining different therapeutic effects and degradation mechanisms in a single molecule.
