Abstract
Dormancy, a reversible quiescent cellular state characterized by greatly reduced metabolic activity, protects from genetic damage, prolongs survival and is crucial for tissue homeostasis and cellular response to injury or transplantation. Dormant cells have been characterized in many tissues, but their identification, isolation and characterization irrespective of tissue of origin remains elusive. Here, we develop a live cell ratiometric fluorescent Optical Stem Cell Activity Reporter (OSCAR) based on the observation that phosphorylation of RNA Polymerase II (RNApII), a hallmark of active mRNA transcription elongation, is largely absent in dormant stem cells from multiple lineages. Using the small intestinal crypt as a model, OSCAR reveals in real time the dynamics of dormancy induction and cellular differentiation in vitro, and allows the identification and isolation of several populations of transcriptionally diverse OSCARhigh and OSCARlow intestinal epithelial cell states in vivo. In particular, this reporter is able to identify a dormant OSCARhigh cell population in the small intestine. OSCAR therefore provides a tool for a better understanding of dormant stem cell biology.
Highlights
Dormancy, a reversible quiescent cellular state characterized by greatly reduced metabolic activity, protects from genetic damage, prolongs survival and is crucial for tissue homeostasis and cellular response to injury or transplantation
A reliable, specific and conserved marker of dormant somatic stem cells that permits their visualization and isolation remains elusive. Even though it has been known for more than 40 years that dormant cells display a lower amount of total RNA18, the molecular mechanism behind this phenomenon is poorly understood. mRNA is transcribed by RNA polymerase II
Since translation depends on mRNA being produced by transcription we examined several tissues for adult stem cells exhibiting low levels of RNA Polymerase II (RNApII)-Ser[2] phosphorylation, a critical determinant of active transcription elongation (Fig. S1a)
Summary
A reversible quiescent cellular state characterized by greatly reduced metabolic activity, protects from genetic damage, prolongs survival and is crucial for tissue homeostasis and cellular response to injury or transplantation. More recent studies, highlighting the high plasticity of the intestinal crypt cells, showed that the reserve stem pool might be composed by differentiated (or early differentiated cells) that can revert to a Lgr5+ stem state in case of need, for example, the Dll1+, Prox1+, or Krt19+ cells[15,16,17] These data suggest the intestinal stem compartment is highly plastic and composed of numerous potential dormant stem cell populations. Somatic stem cells of multiple lineages display low or absent RNApII Serine 2 phosphorylation (RNApII-pSer2), suggesting inactivity of CDK9/ CCNT1 and global down-regulation of productive mRNA transcription in adult stem cells[24]
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