PGLYRP2 drives hepatocyte-intrinsic innate immunity by trapping and clearing hepatitis B virus
Ying Li, … , Yuanfei Yao, Ming Shi
Ying Li, … , Yuanfei Yao, Ming Shi
Published February 13, 2025
Citation Information: J Clin Invest. 2025;135(8):e188083. https://doi.org/10.1172/JCI188083.
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Research Article Hepatology Virology

PGLYRP2 drives hepatocyte-intrinsic innate immunity by trapping and clearing hepatitis B virus

Abstract

Spontaneous clearance of hepatitis B virus (HBV) is frequent in adults (95%) but rare in infants (5%), emphasizing the critical role of age-related hepatic immunocompetence. However, the underlying mechanisms of hepatocyte-specific immunosurveillance and age-dependent HBV clearance remain unclear. Here, we identified PGLYRP2 as a hepatocyte-specific pattern recognition receptor with age-dependent expression, and demonstrated that phase separation of PGLYRP2 was a critical driver of spontaneous HBV clearance in hepatocytes. Mechanistically, PGLYRP2 recognized and potentially eliminated covalently closed circular DNA via phase separation, coordinated by its intrinsically disordered region and HBV DNA-binding domain (PGLYRP2IDR/209–377) in the nucleus. Additionally, PGLYRP2 suppressed HBV capsid assembly by directly interacting with the viral capsid, mediated by its PGRP domain. This interaction promoted the nucleocytoplasmic translocation of PGLYRP2 and subsequent secretion of the PGLYRP2/HBV capsid complex, thereby bolstering the hepatic antiviral response. Pathogenic variants or deletions in PGLYRP2 impaired its ability to inhibit HBV replication, highlighting its essential role in hepatocyte-intrinsic immunity. These findings suggest that targeting the PGLYRP2-mediated host-virus interaction may offer a potential therapeutic strategy for the development of anti-HBV treatments, representing a promising avenue for achieving a functional cure for HBV infection.

Authors

Ying Li, Huihui Ma, Yongjian Zhang, Tinghui He, Binyang Li, Haoran Ren, Jia Feng, Jie Sheng, Kai Li, Yu Qian, Yunfeng Wang, Haoran Zhao, Jie He, Huicheng Li, Hongjin Wu, Yuanfei Yao, Ming Shi

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Figure 3

Role of PGLYRP2IDR-mediated phase separation in trapping HBV DNA and impact of pathogenic variants in human PGLYRP2209–377 on viral inhibition.

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Role of PGLYRP2IDR-mediated phase separation in trapping HBV DNA and imp...
(A–C) Schematic representation and functional assays of PGLYRP2 variants. (A) Top: Schematic representation of full-length (FL) and truncated forms of PGLYRP2. Bottom: Cotransfection assays in C3A cells used HBV promoter–luciferase reporter constructs with either FL or truncated PGLYRP2 plasmids, alongside HBV or HBc expression constructs. (B) ELISA and PCR analysis in stable HepAD38 cell lines for supernatant levels of HBsAg and HBeAg. (C) Real-time PCR determined the levels of intracellular HBV DNA. (D) Prediction of intrinsically disordered regions in PGLYRP2 using the PONDR tool. (E) Expression and phase separation analysis of DsRed-tagged PGLYRP2IDR/209–377 and DsRed-tagged PGLYRP2IDR. DsRed-PGLYRP2IDR/209–377 facilitated the formation of membraneless condensates that colocalized with HBV DNA FAM–Enh II. Scale bars: 10 μm. (F) FRAP analysis of HBV DNA FAM–Enh II within PGLYRP2IDR/209–377-induced condensates. Right: Quantification of fluorescence intensity. Scale bars: 5 μm. (G) Model of PGLYRP2-mediated phase separation as a platform for trapping HBV DNA. (H) Relative luciferase activity analysis of PGLYRP2 variants with missense SNPs. (I) Structural and protein-DNA docking analyses. Conformations of PGLYRP2 and HBV DNA Enh II were predicted using AlphaFold3 and UNAFold, respectively. Protein-DNA docking with HDOCK elucidated specific interactions between the bent structure of Enh II and the PGLYRP2209–377 pocket. (J) HBV promoter DNA pull-down assay. Cell lysates from HEK293 cells transfected with FL PGLYRP2, WT, or SNP-containing PGLYRP2209–377 constructs were incubated with 5′ biotinylated HBV promoter DNA probes or control DNA and streptavidin-agarose beads. Data are represented as mean ± SD. One-way ANOVA with post hoc Bonferroni’s test was used for statistical analysis (A–C and H). *P < 0.05; **P < 0.001.