ATF4 suppresses hepatocarcinogenesis by inducing SLC7A11 (xCT) to block stress-related ferroptosis

Abstract

Hepatocellular carcinoma (HCC), a leading cause of cancer death, is associated with viral hepatitis, non-alcoholic and alcoholic steatohepatitis (NASH, ASH), all of which trigger endoplasmic reticulum (ER) stress, hepatocyte death, inflammation, and compensatory proliferation. Using ER stress-prone MUP-uPA mice, we established that ER stress and hypernutrition cooperate to cause NASH and HCC, but the contribution of individual stress effectors, such as ATF4, to HCC and their underlying mechanisms of action remained unknown. Hepatocyte-specific ATF4 deficient MUP-uPA mice (MUP-uPA/Atf4Δhep) and control MUP-uPA/Atf4F/F mice were fed high fat diet (HFD) to induce NASH-induced HCC, and Atf4F/F and Atf4Δhep mice were injected with diethylnitrosamine (DEN) to model carcinogen-induced HCC. Histological, biochemical, and RNA sequencing analyses were performed to identify and define the role of ATF4-induced SLC7A11 expression in hepatocarcinogenesis. Reconstitution of SLC7A11 in ATF4-deficient primary hepatocytes and mouse livers was used to study its effects on ferroptosis and HCC development. Hepatocyte ATF4 ablation inhibited hepatosteatosis, but increased susceptibility to ferroptosis, resulting in accelerated HCC development. Although ATF4 activates numerous genes, ferroptosis susceptibility and hepatocarcinogenesis were reversed by ectopic expression of a single ATF4 target, Slc7a11, coding for a subunit of the cystine-glutamate antiporter xCT, which is needed for glutathione (GSH) synthesis. A ferroptosis inhibitor also reduced liver damage and inflammation. ATF4 and SLC7A11 amounts were positively correlated in human HCC and livers of NASH patients. Despite ATF4 being upregulated in established HCC, it serves an important protective function in normal hepatocytes. By maintaining glutathione production ATF4 inhibits ferroptosis-dependent inflammatory cell death, which is known to promote compensatory proliferation and hepatocarcinogenesis. Ferroptosis inhibitors or ATF4 activators may also blunt HCC onset.