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Abstract：
Wnt/β-catenin signals are important regulators of embryonic and adult stem cell self-renewal and differentiation and play causative roles in tumorigenesis. Purified recombinant Wnt3a protein, or Wnt3a-conditioned culture medium, has been widely used to study canonical Wnt signaling in vitro or ex vivo. To study the role of Wnt3a in embryogenesis and cancer models, we developed a Cre recombinase activatable Rosa26(Wnt3a) allele, in which a Wnt3a cDNA was inserted into the Rosa26 locus to allow for conditional, spatiotemporally defined expression of Wnt3a ligand for gain-of-function (GOF) studies in mice. To validate this reagent, we ectopically overexpressed Wnt3a in early embryonic progenitors using the T-Cre transgene. This resulted in up-regulated expression of a β-catenin/Tcf-Lef reporter and of the universal Wnt/β-catenin pathway target genes, Axin2 and Sp5. Importantly, T-Cre; Rosa26(Wnt3a) mutants have expanded presomitic mesoderm (PSM) and compromised somitogenesis and closely resemble previously studied T-Cre; Ctnnb1(ex3) (β-catenin(GOF) ) mutants. These data indicate that the exogenously expressed Wnt3a stimulates the Wnt/β-catenin signaling pathway, as expected. The Rosa26(Wnt3a) mouse line should prove to be an invaluable tool to study the function of Wnt3a in vivo.
摘要：
Wnt/β-catenin信号是胚胎和成体干细胞自我更新和分化的重要调节因子，在肿瘤发生中起着重要的致病作用。纯化的重组Wnt3a蛋白,或Wnt3a条件培养基，已广泛用于体外或离体研究经典Wnt信号传导。研究Wnt3a在胚胎发生和癌症模型中的作用，我们开发了Cre重组酶可激活的Rosa26（Wnt3a）等位基因，其中将Wnt3acDNA插入Rosa26基因座以允许有条件的，用于小鼠功能获得(GOF)研究的Wnt3a配体的时空定义表达。为了验证这个试剂,我们使用T-Cre转基因在早期胚胎祖细胞中异位过表达Wnt3a。这导致β-catenin/Tcf-Lef报告基因和通用Wnt/β-catenin途径靶基因的表达上调，Axin2和Sp5。重要的是,T-Cre;Rosa26（Wnt3a）突变体具有扩大的前体性中胚层（PSM）和受损的体发生，并且与先前研究的T-Cre非常相似。Ctnnb1（ex3）（β-catenin（GOF））突变体。这些数据表明，外源表达的Wnt3a刺激Wnt/β-catenin信号通路，如预期。Rosa26（Wnt3a）小鼠品系应被证明是研究Wnt3a体内功能的宝贵工具。
