Downregulation of SOCS1 increases interferon-induced ISGylation during differentiation of induced-pluripotent stem cells to hepatocytes

Background & Aims: Increased expression of IFN-stimulated gene 15 (ISG15) and subsequently increased ISGylation are key factors in the host response to viral infection. In this study, we sought to characterize the expression of ISG15, ISGylation, and associated enzymes at each stage of differentiation from induced pluripotent stem cells (iPSCs) to hepatocytes. Methods: To study the regulation of ISGylation, we utilized patient samples and in vitro cell culture models including iPSCs, hepatocytes-like cells, immortalized cell lines, and primary human hepatocytes. Protein/mRNA expression were measured following treatment with poly(I:C), IFNα and HCV infection. Results: When compared to HLCs, we observed several novel aspects of the ISGylation pathway in iPSCs. These include a lower baseline expression of the ISGylation-activating enzyme, UBE1L, a lack of IFN-induced expression of the ISGylation-conjugation enzyme UBE2L6, an attenuated activation of the transcription factor STAT1 and constitutive expression of SOCS1. ISGylation was observed in iPSCs following downregulation of SOCS1, which facilitated STAT1 activation and subsequently increased expression of UBE2L6. Intriguingly, HCV permissive transformed hepatoma cell lines demonstrated higher intrinsic expression of SOCS1 and weaker ISGylation following IFN treatment. SOCS1 downregulation in HCV-infected Huh 7.5.1 cells led to increased ISGylation. Conclusions: Herein, we show that high basal levels of SOCS1 inhibit STAT1 activation and subsequently IFN-induced UBE2L6 and ISGylation in iPSCs. Furthermore, as iPSCs differentiate into hepatocytes, epigenetic mechanisms regulate ISGylation by modifying UBE1L and SOCS1 expression levels. Overall, this study demonstrates that the development of cell-intrinsic innate immunity during the differentiation of iPSCs to hepatocytes provides insight into cell type-specific regulation of host defense responses and related oncogenic processes. Impact and implications: To elucidate the mechanism underlying regulation of ISGylation, a key process in the innate immune response, we studied changes in ISGylation-associated genes at the different stages of differentiation from iPSCs to hepatocytes. We found that high basal levels of SOCS1 inhibit STAT1 activation and subsequently IFN-induced UBE2L6 and ISGylation in iPSCs. Importantly, epigenetic regulation of SOCS1 and subsequently ISGylation may be important factors in the development of cell type-specific host defense responses in hepatocytes that should be considered when studying chronic infections and oncogenic processes in the liver.