Molecular characterisation of a mutant kras-driven zebrafish model of hepatocellular carcinoma

We used doxycycline (dox) treatment to generate a larval model of hepatocellular carcinoma (HCC)22. In this model, we induced the expression of a single EGFP-krasG12V transgene, denoted TO(krasG12V)T/+ in developing livers by treating with doxycycline between 2-7 days post-fertilisation (dpf). This led to the accumulation of a constitutively active, EGFP-tagged, potently oncogenic form of Kras specifically in hepatocytes, causing hepatocyte hyperplasia and a substantial increase in total liver volume. We used RNA sequencing to analyse the gene expression patterns of hepatocytes expressing the krasG12V transgene compared to wildtype (WT) livers expressing no transgene. We detected more than 6000 significantly upregulated genes in dox-treated TO(krasG12V)T/+ livers compared WT livers, and a further 6000+ genes were significantly downregulated. Of the upregulated genes, many were significantly enriched in KEGG pathways associated with highly proliferative cancers, including DNA replication, cell cycle and DNA damage repair. Geneset enrichment analysis identified a positive correlation between the differential gene expression data from the dox-treated TO(krasG12V)T/+ versus WT livers and the differential gene expression data for the HCC (LIHC) and healthy liver subsets available from The Cancer Genome Atlas. We also found a positive correlation between the DEGs from the dox-treated TO(krasG12V)T/+ versus WT livers and a small HCC expression signature based on four patient samples carrying KRAS G12 or KRAS G13 mutations. These observations build on previous reports that dox-treated TO(krasG12V)T/+ zebrafish provide an authentic model of human HCC. Paired-end RNA-sequencing data was generated for 8 samples of zebrafish liver. Of these samples, 3 are wild type (WT) and 5 are kras mutants. Differential analyses were performed, identifying differentially expressed genes between the kras mutant and wild type groups. Please note, all WT and 2 of the mutant samples are sequenced over 2 runs.