Evaluating the landscape of gene cooperativity with receptor tyrosine kinases in liver tumorigenesis using transposon-mediated mutagenesis

Yannan Fan, Sehrish K. Bazai, Fabrice Daian, Maria Arechederra, Sylvie Richelme, Nuri A. Temiz, Annie Yim, Bianca H. Habermann, Rosanna Dono, David A. Largaespada, Flavio Maina

Research output: Contribution to journalArticlepeer-review

6 Scopus citations

Abstract

Background & Aims: The variety of alterations found in hepatocellular carcinoma (HCC) makes the identification of functionally relevant genes and their combinatorial actions in tumorigenesis challenging. Deregulation of receptor tyrosine kinases (RTKs) is frequent in HCC, yet little is known about the molecular events that cooperate with RTKs and whether these cooperative events play an active role at the root of liver tumorigenesis. Methods: A forward genetic screen was performed using Sleeping Beauty transposon insertional mutagenesis to accelerate liver tumour formation in a genetic context in which subtly increased MET RTK levels predispose mice to tumorigenesis. Systematic sequencing of tumours identified common transposon insertion sites, thus uncovering putative RTK cooperators for liver cancer. Bioinformatic analyses were applied to transposon outcomes and human HCC datasets. In vitro and in vivo (through xenografts) functional screens were performed to assess the relevance of distinct cooperative modes to the tumorigenic properties conferred by RTKs. Results: We identified 275 genes, most of which are altered in patients with HCC. Unexpectedly, these genes are not restricted to a small set of pathway/cellular processes, but cover a large spectrum of cellular functions, including signalling, metabolism, chromatin remodelling, mRNA degradation, proteasome, ubiquitination, cell cycle regulation, and chromatid segregation. We validated 15 tumour suppressor candidates, as shRNA-mediated targeting confers tumorigenicity to RTK-sensitized cells, but not to cells with basal RTK levels. This demonstrates that the context of enhanced RTK levels is essential for their action in tumour initiation. Conclusion: Our study identifies unanticipated genetic interactions underlying gene cooperativity with RTKs in HCC. Moreover, these results show how subtly increased levels of wild-type RTKs provide a tumour permissive cellular environment allowing a large spectrum of deregulated mechanisms to initiate liver cancer. Lay summary: Receptor tyrosine kinases (RTKs) are among signals frequently deregulated in patients with hepatocellular carcinoma and their deregulation confers essential biological properties to cancer cells. We have applied a genetic method to randomly mutate large numbers of genes in the context of a mouse model with increased RTK levels, predisposed to develop liver cancer. We identified mechanisms that accelerate tumour formation in cooperation with enhanced RTK levels. The wide array of cellular functions among these cooperators illustrates an extraordinary capability of RTKs to render the liver more vulnerable to additional alterations, by priming cells for tumour initiation.

Original languageEnglish (US)
Pages (from-to)470-482
Number of pages13
JournalJournal of Hepatology
Volume70
Issue number3
DOIs
StatePublished - Mar 2019

Bibliographical note

Funding Information:
This work was funded by INCa (Institut National du Cancer; PL06_078 and PLBIO12-057), FdF (Fondation de France; 2014_00051580 and 2016_00067080), ARC (Association pour la Recherche contre le Cancer; SFE2011_1203807), and GEFLUC ? Les Entreprises contre le Cancer to F.M. Y.F. was supported by the China Scholarship Council (201206350070). S.K.B. was supported by the Higher Education Commission (HEC) of Pakistan ? France Campus. M.A. was supported by a FdF fellowship. D.A.L. was supported by an American Cancer Society Research Professorship. AY was supported by grant HA 6905/2-1 of the Deutsche Forschungsgemeinschaft (DFG) and an ERASMUS+ fellowship. The contribution of the Region Provence Alpes C?tes d'Azur and of the Aix-Marseille Universit? to the IBDM animal facility is also acknowledged. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Funding Information:
This work was funded by INCa (Institut National du Cancer; PL06_078 and PLBIO12-057), FdF (Fondation de France; 2014_00051580 and 2016_00067080), ARC (Association pour la Recherche contre le Cancer; SFE2011_1203807), and GEFLUC – Les Entreprises contre le Cancer to F.M. Y.F. was supported by the China Scholarship Council (201206350070). S.K.B. was supported by the Higher Education Commission (HEC) of Pakistan – France Campus. M.A. was supported by a FdF fellowship. D.A.L. was supported by an American Cancer Society Research Professorship. AY was supported by grant HA 6905/2-1 of the Deutsche Forschungsgemeinschaft (DFG) and an ERASMUS+ fellowship. The contribution of the Region Provence Alpes Côtes d'Azur and of the Aix-Marseille Université to the IBDM animal facility is also acknowledged. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Funding Information:
This work was funded by INCa (Institut National du Cancer; PL06_078 and PLBIO12-057 ), FdF (Fondation de France; 2014_00051580 and 2016_00067080 ), ARC (Association pour la Recherche contre le Cancer; SFE2011_1203807 ), and GEFLUC – Les Entreprises contre le Cancer to F.M. Y.F. was supported by the China Scholarship Council (201206350070 ). S.K.B. was supported by the Higher Education Commission (HEC) of Pakistan – France Campus. M.A. was supported by a FdF fellowship. D.A.L. was supported by an American Cancer Society Research Professorship . AY was supported by grant HA 6905/2-1 of the Deutsche Forschungsgemeinschaft (DFG) and an ERASMUS+ fellowship. The contribution of the Region Provence Alpes Côtes d’Azur and of the Aix-Marseille Université to the IBDM animal facility is also acknowledged. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Keywords

  • Functional screening
  • HCC
  • Hepatocellular carcinoma
  • Liver cancer
  • Mouse model
  • Oncogenes
  • Receptor tyrosine kinases
  • Signalling
  • Tumour suppressors

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