A novelAxin2knock-in mouse model for visualization and lineage tracing of WNT/CTNNB1 responsive cells

Abstract

2 These authors contributed equally 3 Correspondence to: r.vanamerongen@uva.nl Twitter: @wntlab WNT off Axin2 tetO-Tg GSK3 CTNNB1 WNT on Axin2 rtTA3 tetO-Tg CTNNB1 C T N N B 1 + DOX Axin2 rtTA3 tetO-Tg C T N N B 1 WNT onA novel mouse model to track and trace WNT/CTNNB1 responsive cells 2 Abstract Wnt signal transduction controls tissue morphogenesis, maintenance and regeneration in all multicellular animals. In mammals, the WNT/CTNNB1 (Wnt/b-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 doxycyclineinducible driver for lineage tracing (rtTA3). We show that the Axin2 P2A-rtTA3-T2A-3xNLS-SGFP2 strain labels WNT/CTNNB1 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.

Identification of bipotent progenitors that give rise to myogenic and connective tissues in mouse

“…It is thus tempting to speculate that in the absence of neural crest, mesoderm-derived progenitors can give rise to connective tissue by maintaining canonical Wnt activity. To test this hypothesis, we examined the expression of Axin2, a common readout for Wnt/β-cat activity ( Babb et al, 2017 ; van de Moosdijk et al, 2020 ). Interestingly, Axin2 levels were elevated in the non-myogenic portion of all the different datasets ( Figure 7—figure supplement 1E-H ).…”

Essentiality of CTNNB1 in Malignant Transformation of Human Embryonic Stem Cells under Long-Term Suboptimal Conditions

“…In mammals, CTNNB1 controls cell proliferation and cell fate decisions before and after birth and plays a crucial role at multiple stages of embryonic development [ 34 ]. The current results also indicated that in normal physiological conditions, CTNNB1 played an essential role in maintaining cell proliferation, migration, and differentiation capacities of hESCs; it also participates in cell cycle and telomere regulation.…”

Cell Signaling Pathway Reporters in Adult Hematopoietic Stem Cells

“…Lastly, the group of Roel Nusse, created Wnt reporters with cell lineage tracing capacity by adopting an Axin2 responsive element with a Cre-recombining reporter cassette in the Rosa26 locus [ 19 ]. The most recent Wnt reporter, the Axin2 P2A-rtTA3-T2A-3xNLS-SGFP2 murine model, makes use of a knock-in multi-cystronic cassette upstream of the Axin2 stop codon containing an rtTA3 doxycycline-inducible driver, a triple nuclear localization signal GFP fusion protein and self-cleaving 2A peptides, which preserves both functional alleles of the Axin2 gene [ 20 ]. Notwithstanding the elegant design, this Axin2 reporter detection also suffers from lowly detectable reporter signaling as all fluorescent protein Axin2 reporter models and even hinders the full potential of their employed Wnt driven tetO-Cre strategy for lineage tracing.…”