A single-cell RNA sequence of crypt cells isolated from the adult mouse small intestine

Under homeostatic conditions the intestinal epithelium is replaced daily, mainly via the proliferation of Lgr5+ crypt-base columnar cells (CBCs). Upon intestinal CBC damage, recovery and regeneration are thought to be taken up by quiescent reserve intestinal stem cells (RSCs). However, markers of RSCs have been suggested, but tend to lack specificity. Our previous studies demonstrated an evolutionarily conserved mechanism of cellular quiescence/dormancy, epigenetically regulated by Set domain-containing protein 4 (Setd4) and maintained via asymmetric division. To investigate whether a subgroup of cells underpin this Setd4 signature, we performed single-cell RNA sequence of crypt cells (7-AAD-CD45-EpCAM+CD44+) isolated from the adult mouse small intestine using fluorescence-activated cell sorting (FACS). The previously proposed +4/RSC markers Bmi12(18.8%), Hopx13 (61.3%) and Lrig111(31.7%) were expressed in several clusters, whereas Mex3a14 (6.9%) was highly expressed in a few scattered cells and Tert10 was not detected. Contrastingly, Setd4+ cells represented 3.6% of crypt cells, but at a particularly high level (86.1%) across RSC populations and at a lower level (26.2%) across Lgr5+ CBC populations. Thus, we re-identified a small relatively quiescent population of cells in the intestines of adult mice with RSC characteristics.