Hepatitis B virus (HBV) core protein (HBc) plays many roles in the HBV life cycle, such as regulation of transcription, RNA encapsidation, reverse transcription, and viral release. To accomplish these functions, HBc interacts with many host proteins and undergoes different post-translational modifications (PTMs). One of the most common PTMs is ubiquitination, which was shown to change the function, stability, and intracellular localization of different viral proteins, but the role of HBc ubiquitination in the HBV life cycle remains unknown. Here, we found that HBc protein is post-translationally modified through K29-linked ubiquitination. We performed a series of co-immunoprecipitation experiments with wild-type HBc, lysine to arginine HBc mutants and wild-type ubiquitin, single lysine to arginine ubiquitin mutants, or single ubiquitin-accepting lysine constructs. We observed that HBc protein could be modified by ubiquitination in transfected as well as infected hepatoma cells. In addition, ubiquitination predominantly occurred on HBc lysine 7 and the preferred ubiquitin chain linkage was through ubiquitin-K29. Mass spectrometry (MS) analyses detected ubiquitin protein ligase E3 component N-recognin 5 (UBR5) as a potential E3 ubiquitin ligase involved in K29-linked ubiquitination. These findings emphasize that ubiquitination of HBc may play an important role in HBV life cycle.

• Klíčová slova
• E3 ubiquitin-protein ligase, HBc, hepatitis B virus, post-translational modifications, ubiquitin, ubiquitination,
• MeSH
• arginin genetika   MeSH
• buněčné linie MeSH
• buňky Hep G2 MeSH
• HEK293 buňky MeSH
• hepatitida B genetika   MeSH
• hepatocelulární karcinom genetika   MeSH
• lidé MeSH
• lysin genetika   MeSH
• nádorové buněčné linie MeSH
• posttranslační úpravy proteinů genetika   MeSH
• ubikvitin genetika   MeSH
• ubikvitinace genetika   MeSH
• ubikvitinligasy genetika   MeSH
• virové proteiny genetika   MeSH
• virus hepatitidy B genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• arginin MeSH
• lysin MeSH
• ubikvitin MeSH
• ubikvitinligasy MeSH
• virové proteiny MeSH

In mammals, protein arginine methyltransferase 5, PRMT5, is the main type II enzyme responsible for the majority of symmetric dimethylarginine formation in polypeptides. Recent study reported that PRMT5 restricts Hepatitis B virus (HBV) replication through epigenetic repression of HBV DNA transcription and interference with encapsidation of pregenomic RNA. Here we demonstrate that PRMT5 interacts with the HBV core (HBc) protein and dimethylates arginine residues within the arginine-rich domain (ARD) of the carboxyl-terminus. ARD consists of four arginine rich subdomains, ARDI, ARDII, ARDIII and ARDIV. Mutation analysis of ARDs revealed that arginine methylation of HBc required the wild-type status of both ARDI and ARDII. Mass spectrometry analysis of HBc identified multiple potential ubiquitination, methylation and phosphorylation sites, out of which lysine K7 and arginines R150 (within ARDI) and R156 (outside ARDs) were shown to be modified by ubiquitination and methylation, respectively. The HBc symmetric dimethylation appeared to be linked to serine phosphorylation and nuclear import of HBc protein. Conversely, the monomethylated HBc retained in the cytoplasm. Thus, overexpression of PRMT5 led to increased nuclear accumulation of HBc, and vice versa, down-regulation of PRMT5 resulted in reduced levels of HBc in nuclei of transfected cells. In summary, we identified PRMT5 as a potent controller of HBc cell trafficking and function and described two novel types of HBc post-translational modifications (PTMs), arginine methylation and ubiquitination.

• MeSH
• fosforylace MeSH
• hmotnostní spektrometrie MeSH
• lidé MeSH
• metylace MeSH
• proteinarginin-N-methyltransferasy metabolismus   fyziologie   MeSH
• replikace viru fyziologie   MeSH
• subcelulární frakce metabolismus   MeSH
• ubikvitinace MeSH
• virus hepatitidy B enzymologie   fyziologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• PRMT5 protein, human MeSH Prohlížeč
• proteinarginin-N-methyltransferasy MeSH