The impact of HBx protein on mitochondrial dynamics and associated signaling pathways strongly depends on the hepatitis B virus genotype

Abstract

ABSTRACT Chronic hepatitis B virus (HBV) infections are strongly associated with liver cirrhosis, inflammation, and hepatocellular carcinoma. In this context, the viral HBx protein is considered as a major factor influencing HBV-associated pathogenesis through deregulation of multiple cellular signaling pathways and is therefore a potential target for prognostic and therapeutic applications. However, HBV-associated pathogenesis differs significantly between genotypes, with the relevant factors and in particular the contribution of the genetic diversity of HBx being largely unknown. To address this question, we studied the specific genotype-dependent impact of HBx on cellular signaling pathways, focusing in particular on morphological and functional parameters of mitochondria. To exclusively investigate the impact of HBx of different genotypes on integrity and function of mitochondria in the absence of additional viral factors, we overexpressed HBx in Huh7 or HepG2 cells. Key signaling pathways were profiled by kinome analysis and correlated with expression levels of mitochondrial and pathogenic markers. Conclusively, HBx of genotypes A and G caused strong disruption of mitochondrial morphology alongside an induction of PTEN-induced putative kinase 1/Parkin-mediated mitophagy. These effects were only moderately dysregulated by genotypes B and E, whereas genotypes C and D exhibit an intermediate effect in this regard. Accordingly, changes in mitochondrial membrane potential and elevated reactive oxygen species production were associated with the HBx-mediated dysfunction among different genotypes. Also, genotype-related differences in mitophagy induction were identified and indicated that HBx-mediated changes in the mitochondria morphology and function strongly depend on the genotype. This indicates a relevant role of HBx in the process of genotype-dependent liver pathogenesis of HBV infections and reveals underlying mechanisms. IMPORTANCE The hepatitis B virus is the main cause of chronic liver disease worldwide and differs in terms of pathogenesis and clinical outcome among the different genotypes. Furthermore, the viral HBx protein is a known factor in the progression of liver injury by inducing aberrant mitochondrial structures and functions. Consequently, the selective removal of dysfunctional mitochondria is essential to maintain overall cellular homeostasis and cell survival. Consistent with the intergenotypic difference of HBV, our data reveal significant differences regarding the impact of HBx of different genotypes on mitochondrial dynamic and function and thereby on radical oxygen stress levels within the cell. We subsequently observed that the induction of mitophagy differs significantly across the heterogenetic HBx proteins. Therefore, this study provides evidence that HBx-mediated changes in the mitochondria dynamics and functionality strongly depend on the genotype of HBx. This highlights an important contribution of HBx in the process of genotype-dependent liver pathogenesis.