RNA Seq analysis of Schmidtea mediterranea treated with RNAi against chd4, p53, or unc22 to identify factors involved in neoblast differentiation.. Schmidtea mediterranea

Adult stem cells are tissue-specific cells with the capacity to self-renew and differentiate to continually replace cells lost to normal physiological turnover or injury. Neoblasts, the planarian stem cells, are widely distributed throughout the body mesenchyme, driving constitutive renewal of tissues during homeostasis and endowing planarians with the remarkable capacity to regenerate wholly from tiny tissue fragments. Neoblasts are the only dividing cells in planarians and are believed to be collectively comprised of both a heterogeneous population of pluripotent cells with broad differentiation potential and also lineage-committed progenitor cells that give rise to specific tissues. Recent work has identified a prominent class of neoblasts referred to as zeta-class, which express the zinc-finger gene zfp-1. Zfp-1, along with the chromatin remodeling factor chd4, the tumor suppressor gene p53, and most recently an RNA-binding protein mex3-1, have all been shown to be required for maintenance of two related post-mitotic, sub-epidermal cell populations expressing the specific marker genes prog-1 and AGAT-1, which have been widely used to asses neoblast differentiation. Given that zeta-class neoblasts are required for the generation of prog-1 and AGAT-1 positive cells and other markers of epidermal cell types, prog-1 and AGAT-1 likely mark two major populations of epidermal progeny cells. To understand the molecular mechanisms underlying neoblast differentiation and how they give rise to multiple cell types, we devised a strategy to identify critical factors enriched in the AGAT-1 positive cell population required for epidermal lineage progression. We performed RNA-Seq analysis of chd4 and p53 RNAi animals and characterized additional markers enriched in AGAT-1 positive and related post-mitotic cells. Overall design: Examine gene expression profiles of adult flatworms after 3, 6, 9, 12, 15, or 18 days of treatment with RNAi against either chd4, p53, or an unc22 non-targeting control. Experiments were performed in triplicate yielding a total of 50 samples after removal of an outlier. The chd4 time course extends to only 15 days due to lethality.