A novel Axin2 knock‐in mouse model for visualization and lineage tracing of WNT/CTNNB1 responsive cells

Abstract

Summary Wnt signal transduction controls tissue morphogenesis, maintenance and regeneration in all multicellular animals. In mammals, the WNT/CTNNB1 (Wnt/β‐catenin) pathway controls cell proliferation and cell fate decisions before and after birth. It plays a critical role at multiple stages of embryonic development, but also governs stem cell maintenance and homeostasis in adult tissues. However, it remains challenging to monitor endogenous WNT/CTNNB1 signaling dynamics in vivo. Here, we report the generation and characterization of a new knock‐in mouse strain that doubles as a fluorescent reporter and lineage tracing driver for WNT/CTNNB1 responsive cells. We introduced a multi‐cistronic targeting cassette at the 3′ end of the universal WNT/CTNNB1 target gene Axin2. The resulting knock‐in allele expresses a bright fluorescent reporter (3xNLS‐SGFP2) and a doxycycline‐inducible driver for lineage tracing (rtTA3). We show that the Axin2P2A‐rtTA3‐T2A‐3xNLS‐SGFP2 strain labels WNT/CTNNB1 responsive cells at multiple anatomical sites during different stages of embryonic and postnatal development. It faithfully reports the subtle and dynamic changes in physiological WNT/CTNNB1 signaling activity that occur in vivo. We expect this mouse strain to be a useful resource for biologists who want to track and trace the location and developmental fate of WNT/CTNNB1 responsive stem cells in different contexts.

Wnt/β-Catenin Signaling Promotes Differentiation of Ischemia-Activated Adult Neural Stem/Progenitor Cells to Neuronal Precursors

“…However, subsequent analysis of expression profiles at the single-cell level provided a slightly more “prosaic” explanation. Like other evolutionarily conserved signaling pathways, the Wnt pathway does not always regulate the same group of genes and the transcriptional response to its activation (or inhibition) is thus tissue-specific; although there is a consensus that Axin2 represents such a “universal” target gene ( van de Moosdijk et al, 2020 ). The analysis of the expression levels of selected genes ( Axin2 , Nkd1 , and Troy ) in individual cell subpopulations clearly showed their different expression profile ( Figure 7 ).…”

The Wnt Pathway: From Signaling Mechanisms to Synthetic Modulators

“…Wnt transcription activity can be measured through reporters such as TOPFLASH, which contains multimerized TCF/LEF-binding motifs (250, 251). Negative feedback regulators of Wnt signaling such as AXIN2 are general Wnt target genes and serve as a faithful readout of Wnt pathway activation through mRNA quantification or visualization of AXIN2-driven reporters (38,(252)(253)(254)(255).…”

Evidence for lung barrier regeneration by differentiation prior to binucleated and stem cell division

“…Conditional fluorescent Cre reporter alleles included: R26H2B-EGFP ( Abe et al, 2011 ; CBD0238K; RIKEN); R26Tomato ( Madisen et al, 2010 ; #007914; JAX Strain); R26mTmG ( Muzumdar et al, 2007 ; #007676; JAX Strain), in which recombination switches expression from membrane-targeted Tomato to mGFP; and the R26iMb-Mosaic ( Pontes-Quero et al, 2017 ; #031298; JAX Strain) allele, in which recombination induces stochastic expression of either membrane-targeted YFP, Tomato, or Kate2. Doxycycline-induced (1 mg/ml doxycycline in 1% sucrose drinking water; Abcam) fluorescent cell nuclear labeling was performed using Sftpc-CreER-rtTA with Col1a1TetoH2BCherry ( Egli et al, 2007 ; #014592; JAX Strain) for AT2 cells and Axin2-rtTA-nls-GFP ( van de Moosdijk et al, 2020 ; #036013; JAX Strain) with tetO-H2B-GFP ( Tumbar et al, 2004 ; #005104; JAX Strain) for Wnt/β-catenin–responsive cells. Inducible, gene-targeted cell ablation employed the R26iDTR ( Buch et al, 2005 ; #007900; JAX Strain) allele, which sensitizes cells to DT by conditional expression of the toxin receptor, and the R26DTA ( Voehringer et al, 2008 ; #009669; JAX Strain) allele, which kills cells by conditionally inducing synthesis of the cytotoxic protein fragment.…”