Characterization of PRMT associated with the NS3 of hepatitis C virus

Abstract

The Hepatitis C virus (HCV) is a major inducer of chronic hepatitis, and its chronic infection is associated with liver cirrhosis, and eventually, development of hepatocellular carcinoma. To develop therapeutic tools against of hepatocellular carcinoma caused by chronic infection of HCV, it is important to understand molecular mechanism by which HCV causes liver pathogenesis and which cellular genes are involved in hepatocellular carcinogenesis. A number of studies suggest that the core protein of HCV is a major viral factor in in hepatocellular carcinogenesis. Also, NS3 protein of HCV has been shown to be implicated in the liver pathogenesis. To search for cellular targets of the NS3 protein, we conducted a yeast two-hybrid screen. We found that Janus kinase-binding protein 1 (JBP1), known as a human homologue of Shk1 kinase-binding protein 1 of the fission yeast (Skb1Hs), interacts with the nonstructural protein 3 of the hepatitis C virus by a yeast two-hybrid screen. Amino acid sequence analysis revealed that Skb1Hs/JBP1 contains conserved motifs of S-adenosylmethionine-dependent protein arginine methyltransferases (PRMTs). Here, we demonstrate that Skb1Hs/JBP1, named PRMT5, is a distinct member of the protein arginine methyltransferase family. Recombinant PRMT5 protein purified from human cells methylates myelin basic protein and histone. Myelin basic protein methylated by PRMT5 contains monomethylated and dimethylated arginine residues. Sedimentation analysis of the purified PRMT5 on a sucrose density gradient indicates that PRMT5 forms distinct homo-oligomeric complexes, including dimer and tetramer, which comigrate with the enzyme activity. Western blot analysis of sedimentation fractions suggests that endogenous PRMT5 is present as a homo-oligomer in a 293T cell extract. This finding prompted us to examine the possibility of whether arginine residues of the NS3 protein are methylated by protein arginine methyltransferases (PRMTs). PRMTs transfer the me...