Introduction
Primary liver cancer is the 6th most commonly diagnosed cancer and the 3rd leading cause of cancer death worldwide. Incidence and mortality are higher among men and in low to moderate income countries (1). Hepatocellular carcinoma (HCC), accounting for 75–85% of primary liver cancers, develops in the context of chronic liver diseases, such as hepatitis and/or metabolic dysfunction. HCC is increasingly associated with obesity, insulin resistance and the metabolic syndrome and has limited therapeutic options (2, 3). The signalling pathways most frequently involved in hepatocarcinogenesis include Wnt/β-catenin, mTOR, IL-6, TGF-β, Ras, Rb, HGF/c-Met, and IGF1, which converge and modulate the activity of the NF-κB, p53, Stat3, c-Myc and AP-1 transcription factors (4-6).
The Jun (c-Jun, JunB, JunD) and Fos (c-Fos, FosB, Fra-1, Fra-2) proteins are components of the dimeric Activator Protein-1 (AP-1) transcription factor complex (7). While Jun proteins can form homo- or heterodimers, Fos proteins can only form heterodimers with a Jun protein. The AP-1 dimer combinations that co-exist in a given cell/biological context, together with dimer-specific variation in DNA sequence affinity and/or co-activator/repressor recruitment, determine the target genes that are positively or negatively regulated by AP-1. The AP-1 dimer pool is modulated by various signals, such as growth factors, inflammatory cytokines, mechanical and oxidative stress, and plays important roles in many diseases including cancer (7, 8). In genetically engineered mouse models (GEMMs), liver-specific inactivation of c-Jun revealed its essential role in liver regeneration (9), steatohepatitis (10), hepatocyte survival during acute hepatitis (11), endoplasmic reticulum (ER) stress (12), and liver cancer (13-18). In HCC, c-Jun promotes the survival of diethylnitrosamine (DEN)-induced pre-neoplastic hepatocytes by repressing c-Fos expression (16), while c-Fos is needed for DEN-induced hepatocarcinogenesis when the c-jun gene is intact (19). Furthermore, doxycycline (Dox)-switchable c-Fos expression in adult hepatocytes (c-Foshep) leads to reversible liver inflammation, accumulation of toxic oxysterols and bile acids, activation of the DNA damage response (DDR), premalignant transformation and enhanced DEN-induced HCC (19). The hepatic functions of the other Jun and Fos proteins are less studied, especially in cancer. Hepatocyte-specific JunB inactivation increases liver damage during acute hepatitis, an effect that is largely counteracted by the pro-inflammatory role of JunB in hepatic NK/NKT cells (20). JunD knock-out mice are protected from chemically-induced liver fibrosis (21) and high fat diet (HFD)-induced hepatosteatosis (22). On the other hand, loss of Fra-1 sensitizes, while hepatic Fra-1, but not Fra-2, expression protects from acetaminophen-induced liver damage, an acute liver failure paradigm (23). Interestingly, Fra-1 and Fra-2 play redundant functions in hepatic lipid metabolism: Fra-1 or Fra-2 expression in hepatocytes prevented and could even revert HFD-induced hepatosteatosis by suppressing the transcription of the nuclear receptor PPARγ, a central regulator of lipid metabolism, while single inactivation of either of the two genes had no effect (22, 24). In contrast, c-Fos activated hepatic Pparg transcription, while it suppressed another nuclear receptor LXRα, responsible for increased hepatic cholesterol and oxysterols (19, 22). Thus, Fra-1/2- and c-Fos-containing AP-1 dimers exert antagonistic effects on the pparg2 promoter and lipid handling in the liver. When selected Jun and Fos monomers were tethered by a flexible polypeptide to force specific AP-1 pairing in a “single-chain” approach (25), and expressed in Dox-switchable AP-1hep mice, c-Jun~Fra-2 dimers inhibited, whereas c-Jun~c-Fos, JunB~c-Fos, and JunD~c-Fos dimers activated PPARγ expression and signalling (22, 24).
In this study, we show that hepatic expression of c-Jun~Fra-2 dimers results in spontaneous and reversible HCC formation, while mice expressing Fra-1/2 monomers or c-Jun~Fra-1 dimers remained tumor-free. c-Jun~Fra-2 dimers promote tumorigenesis in murine and human liver cells, in significant part through direct transcriptional activation of c-myc expression. Furthermore, we show that established tumors are largely addicted to c-Jun~Fra-2 and sensitive to JQ-1, a BET bromodomain inhibitor that inhibits c-Myc activity.