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Regulation of Hippo signaling and planar cell polarity via distinct regions of the Fat intracellular domain [ds38k]

ABSTRACT: The large Drosophila protocadherin Fat (Ft) is a receptor for signal transduction pathways that control growth (Hippo signaling), planar cell polarity (PCP), metabolism and the proximodistal patterning of appendages. The intracellular domain (ICD) of Ft is crucial in implementing its biological functions. Six regions of high conservation (named A-F) within the ICD have been identified, as well as distinct regions mediating Hippo pathway activity that have been functionally characterized via overexpression assays. Here, we make targeted deletions of these highly conserved residues and the putative Hippo- and PCP-regulating domains of endogenous Ft using CRISPR/Cas9. Through transcriptomic, developmental, and phenotypic analyses, we show that different regions of Ft contribute uniquely to chromatin dynamics, tissue morphogenesis, PCP and metabolic regulation. We also demonstrate that different regions of Ft regulate Yorkie activity in opposite directions, finely tuning growth and tissue development. Strikingly, conserved regions D and F are key regulators of Ft’s functions- exhibiting opposing effects on wing morphology and Hippo pathway modulation- and conserved region D is the main regulator of PCP.

ORGANISM(S): Drosophila melanogaster

PROVIDER: GSE288256 | GEO | 2025/01/28

REPOSITORIES: GEO

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Project description:A detailed understanding of intestinal stem cell (ISC) self-renewal and differentiation is required for better treatment options for a variety of chronic intestinal diseases, however, current models of ISC lineage hierarchy and segregation are still under debate. Here we report the identification of Lgr5+ ISCs that express Flattop (Fltp), a Wnt/planar cell polarity (PCP) reporter and effector gene. Functional analysis and lineage tracing revealed that Wnt/PCP-activated Fltp+ ISCs are primed towards either the enteroendocrine or Paneth cell lineage in vivo, while retaining self-renewal and multi-lineage capacity in vitro. Surprisingly, canonical Wnt/beta-catenin- and non-canonical Wnt/PCP-activated Lgr5+ ISCs are indistinguishable by the expression of stem-cell signature or secretory lineage-specifying genes, suggesting that lineage priming and cell-cycle exit is triggered at the post-transcriptional level by polarity cues and a switch of canonical to non-canonical Wnt signalling. Pseudotemporal ordering of targeted single-cell gene expression data allowed us to delineate the ISC differentiation path into enteroendocrine and Paneth cells. Strikingly, both lineages are directly recruited from ISCs via unipotent transition states, excluding the existence of formerly predicted bi- or multipotent secretory progenitors. Transitory cells that mature into Paneth cells are defined by label-retention and co-expression of stem cell and secretory lineage genes, indicating that these cells are the previously described Lgr5+ label-retaining cells (LRCs). Taken together, we identified the Wnt/PCP pathway as a new niche signal and polarity cue regulating stem cell fate. Active Wnt/PCP signalling represents one of the earliest events in ISC lineage priming towards the Paneth and enteroendocrine cell fate, preceding lateral inhibition and expression of secretory lineage-specifying genes. Thus, our findings provide a better understanding of the niche signals and redefine the mechanisms underlying ISC lineage hierarchy and segregation. Here we establish the Wnt/Planar cell polarity (PCP) gene Flattop (Fltp) as a unique marker for intestinal LRCs and their distinct secretory fate. We show that Fltp+ cells are characterized by a combined stem cell and secretory gene signature. A subset of Fltp+ cells is classified by label-retention and predominantly locates at position +4, indicative of quiescent stem cells. Strikingly, Fltp+ LRCs are specified by Wnt/PCP signaling in contrast to actively cycling stem cells that rely on the canonical Wnt/β-catenin pathway. In mice with disturbed Wnt/PCP signaling, the differentiation of enteroendocrine cells from LRCs is impaired. These findings establish Fltp as a novel marker for intestinal LRCs and implicate Wnt/PCP signaling in cell-cycle exit and commitment of ISCs to the secretory lineage. Taken together, we not only provide a marker to study LRCs in homeostatic and diseased conditions, but also identify the Wnt/PCP signaling pathway as a therapeutic target for colorectal cancer and metabolic disease.

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Regulation of Hippo signaling and planar cell polarity via distinct regions of the Fat intracellular domain [ds38k]

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