Infection with hepatitis C virus depends on TACSTD2, a regulator of claudin-1 and occludin highly downregulated in hepatocellular carcinoma

Entry of hepatitis C virus (HCV) into hepatocytes is a complex process that involves numerous cellular factors, including the scavenger receptor class B type 1 (SR-B1), the tetraspanin CD81, and the tight junction (TJ) proteins claudin-1 (CLDN1) and occludin (OCLN). Despite expression of all known HCV-entry factors, in vitro models based on hepatoma cell lines do not fully reproduce the in vivo susceptibility of liver cells to primary HCV isolates, implying the existence of additional host factors which are critical for HCV entry and/or replication. Likewise, HCV replication is severely impaired within hepatocellular carcinoma (HCC) tissue in vivo, but the mechanisms responsible for this restriction are presently unknown. Here, we identify tumor-associated calcium signal transducer 2 (TACSTD2), one of the most downregulated genes in primary HCC tissue, as a host factor that interacts with CLDN1 and OCLN and regulates their cellular localization. TACSTD2 gene silencing disrupts the typical linear distribution of CLDN1 and OCLN along the cellular membrane in both hepatoma cells and primary human hepatocytes, recapitulating the pattern observed in vivo in primary HCC tissue. Mechanistic studies suggest that TACSTD2 is involved in the phosphorylation of CLDN1 and OCLN, which is required for their proper cellular localization. Silencing of TACSTD2 dramatically inhibits HCV infection with a pan-genotype effect that occurs at the level of viral entry. Our study identifies TACSTD2 as a novel regulator of two major HCV-entry factors, CLDN1 and OCLN, which is strongly downregulated in malignant hepatocytes. These results provide new insights into the complex process of HCV entry into hepatocytes and may assist in the development of more efficient cellular systems for HCV propagation in vitro.
Author summary Infection with hepatitis C virus (HCV) is associated with a high risk of developing hepatocellular carcinoma (HCC). However, it is unknown whether HCV promotes HCC indirectly, through chronic inflammation, fibrosis and liver regeneration, or directly, in a manner similar to other oncogenic viruses. We recently demonstrated that HCV replication within HCC tissue is severely hampered, and the present study identifies a potential mechanism for this restriction. We found that HCV RNA suppression in the tumor correlates with disruption of the typical linear distribution of two major HCV-entry cofactors, claudin-1 (CLDN1) and occludin (OCLN), which mediate viral entry into hepatocytes. This abnormal pattern was associated with downregulation of tumor-associated calcium signal transducer 2 (TACSTD2), an essential gene for maintaining the proper cellular localization of CLDN1 and OCLN. Thus, this study identifies TACSTD2 as a novel host factor that regulates the cellular localization of two major HCV-entry cofactors. A major highlight of our study is that our in vitro observations recapitulate the in vivo findings in patients with HCC, corroborating their biological relevance. These results shed new light on the complex process of HCV entry and may have relevance for the pathogenesis, treatment and prevention of HCV infection.