Abstract
ABSTRACTUp to 41% of hepatocellular carcinomas (HCCs) result from activating mutations in the CTNNB1 gene encoding β-catenin. HCC-associated CTNNB1 mutations stabilize the β-catenin protein, leading to nuclear and/or cytoplasmic localization of β-catenin and downstream activation of Wnt target genes. In patient HCC samples, β-catenin nuclear and cytoplasmic localization are typically patchy, even among HCC with highly active CTNNB1 mutations. The functional and clinical relevance of this heterogeneity in β-catenin activation are not well understood. To define mechanisms of β-catenin-driven HCC initiation, we generated a Cre-lox system that enabled switching on activated β-catenin in (1) a small number of hepatocytes in early development; or (2) the majority of hepatocytes in later development or adulthood. We discovered that switching on activated β-catenin in a subset of larval hepatocytes was sufficient to drive HCC initiation. To determine the role of Wnt/β-catenin signaling heterogeneity later in hepatocarcinogenesis, we performed RNA-seq analysis of zebrafish β-catenin-driven HCC. At the single-cell level, 2.9% to 15.2% of hepatocytes from zebrafish β-catenin-driven HCC expressed two or more of the Wnt target genes axin2, mtor, glula, myca and wif1, indicating focal activation of Wnt signaling in established tumors. Thus, heterogeneous β-catenin activation drives HCC initiation and persists throughout hepatocarcinogenesis.
Highlights
Liver cancer is the seventh most commonly occurring cancer in the world and the third highest contributor to cancer-related mortality (Ghouri et al, 2017)
This transgenic line consists of CreERT2 cDNA, encoding Cre recombinase fused to the modified ligand-binding domain of the human estrogen receptor (ERT2) (Branda and Dymecki, 2004; Feil et al, 1997; Metzger and Chambon, 2001), downstream of the hepatocyte-specific fabp10a promoter (Her et al, 2003b)
The lox-switch part of our modular system consists of the Tg(fabp10a:loxP-BFP-loxP-Xla.Ctnnb1) line, hereafter referred to as Tg(fabp10a:flox-pt-β-cat), wherein the fabp10a promoter drives expression of activated β-catenin preceded by a blue fluorescent protein (BFP)-STOP cassette flanked by loxP sites (Fig. 1A)
Summary
Liver cancer is the seventh most commonly occurring cancer in the world and the third highest contributor to cancer-related mortality (Ghouri et al, 2017). Hepatocellular carcinoma (HCC) comprises over 90% of all primary liver cancers (Lin et al, 2017). Approved chemotherapeutic strategies against HCC include multi-kinase inhibitors and immune-checkpoint inhibitors. Patient response to these drugs and improvement in survival are marginal (Zhu and Hoshida, 2018). Genome-wide analyses of clinical HCC samples reveal a range of functional molecular aberrations that can. Received 10 September 2019; Accepted 24 September 2019 be grouped into distinct molecular subclasses (Chiang et al, 2008). A substantial subset of HCC (13–41%) is defined by activating mutations in the gene encoding β-catenin (CTNNB1) (Khalaf et al, 2018)
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