Major HBV splice variant encoding a novel protein important for infection

Area of Science:

• Virology
• Molecular Biology
• Hepatitis Research

Background:

• Hepatitis B virus (HBV) expresses numerous spliced RNAs from its pregenomic RNA, with largely unknown functions.
• Previous studies suggested the integration of a C-terminal cysteine-lacking HBc protein (HBc^-Cys) into the HBV capsid.

Purpose of the Study:

• To elucidate the function of HBV spliced RNAs, specifically SP1.
• To investigate the role of the novel HBc^SP1 protein in HBV infection and viral entry.

Main Methods:

• Generation of splicing-deficient HBV mutants, including A487C.
• Infection studies using HepG2-NTCP cells and human hepatocyte chimeric mice (hu-FRG mice).
• SHifter assays and cryo-electron microscopy to detect and analyze viral components and capsid structure.

Main Results:

• Splicing-deficient HBV mutants showed a 100-1,000-fold decrease in infectivity in hu-FRG mice.
• The A487C mutant, lacking SP1 RNA, exhibited significantly impaired viral entry and infectivity.
• Cryo-electron microscopy revealed a locked capsid conformation in the A487C mutant due to a disulfide bond, hindering viral entry.

Conclusions:

• The study confirms and validates the identity and function of the HBc^SP1 protein during HBV infection.
• HBc^SP1, encoded by SP1 RNA, is essential for efficient HBV infectivity by preventing capsid locking and facilitating viral entry.
• HBc^SP1 is present in HBV particles from cell culture and in the serum of patients with chronic hepatitis B.