The ER resident proteins, VAMP-associated protein (VAP) A and B (VAPA and VAPB) have been reported to be necessary for efficient HCV replication, but the specific mechanisms are not well understood. VAP proteins are known to recruit lipid transfer proteins to the ER, including oxysterol binding protein (OSBP), which has been previously shown to be necessary for cholesterol delivery to the HCV replication organelle in exchange for phosphatidylinositol 4-phosphate (PI (4)P). Here we show that VAPA and VAPB are redundant for HCV infection and that dimerization is not required for their function. In addition, we identify the phosphatidylinositol transfer protein Nir2 as an effector of VAPs to support HCV replication. We propose that Nir2 functions to replenish phosphoinositides at the HCV replication organelle to maintain elevated steady-state levels of PI (4)P, which is removed by OSBP. Thus, Nir2, along with VAPs, OSBP and the phosphatidylinositol 4-kinase, complete a cycle of phosphoinositide flow between the ER and viral replication organelles to drive ongoing viral replication.
Hepatitis C Virus (HCV) is known for its ability to modulate phosphoinositide signaling pathways for its replication. Elevated levels of phosphatidylinositol 4-phosphate (PI (4)P) in HCV replication organelles (ROs) recruits lipid transfer proteins (LTPs), like oxysterol- binding protein (OSBP). OSBP exchanges PI (4)P with cholesterol, thus removing PI (4)P from the HCV RO. Here we found that the phosphatidylinositol transfer protein Nir2 acts as an LTP may replenish PI at the HCV RO by interacting with VAMP associated proteins (VAPs), enabling continuous viral replication during chronic infection. Therefore, the coordination of OSBP, Nir2, VAPs complete our understanding of the phosphoinositide cycle between ER and HCV RO.