Ex vivo induced transcriptome changes in a subpopulation of satellite cells revealed by single-cell mRNA sequencing
Susanne C. van den Brink1, Fanny G. Sage1, Ábel Vértesy1, Bastiaan Spanjaard1, Catherine Robin1,2 and Alexander van Oudenaarden1,*
1Hubrecht Institute–KNAW and University Medical Center Utrecht, 3584 CT Utrecht, The Netherlands; 2Department of Cell Biology, University Medical Center Utrecht, 3584 EA Utrecht, The Netherlands
Regeneration of skeletal muscle in adults depends on the activation of otherwise quiescent muscle stem cells, the satellite cells. Recently it became possible to purify satellite cells from murine skeletal muscle allowing a genome-wide characterization of gene expression in these cells. To obtain a detailed view on how gene expression varies between individual quiescent satellite cells, we applied single-cell mRNA sequencing to freshly isolated satellite cells. These experiments revealed two distinct subpopulations that display vastly different expression levels of heat shock proteins (HSPs) and the immediate early genes (IEGs) including Fos and Jun. Unexpectedly, we were not able to validate the subpopulation with high HSP and IEG expression in situ suggesting that expression of these genes is initiated during the satellite cell purification protocol. Even though the high HSP/IEG expressing subpopulation can be relatively small, these genes are expressed at very high levels resulting in a strong, artificial, signal in bulk gene expression studies in which these subpopulations cannot be separated. In light of this knowledge several conclusions based on previously published bulk expression studies warrant reinterpretation. Since similar dissociation procedures are used to isolate cells from other tissues, our findings might also be relevant for work done on other cell types in other tissues. Finally, we demonstrate that this contaminating population of satellite cells can be effectively removed by utilizing the single-cell transcriptome data.
Credits: None available.
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