Hepatitis B Virus Genome Packaging and Reverse Transcription

Introduction

Hepatitis B Virus (HBV) is an infectious liver virus that contains a genome consisting of circular, double-stranded DNA. HBV is a member of a family of viruses that takes RNA genetic information generated during infection, and transcribes it into DNA to package into new viral particles (1). The process of reverse transcription is unique to viruses, as there is no need to generate DNA from RNA anywhere else in biological life. 

For liver cells infected with HBV to form infectious viral particles, the viral DNA produced during this infection must fit inside new viral particles to generate more viruses to infect more cells (1). This process is called packaging, and is required in all viral infections, regardless of the types of cells they infect, or the type of genome that the virus packages its cells with. Viral genomes are not very large compared to more complex life, such as in human cells, but the size of an average viral particle is also much smaller than an animal cell, and fitting the entire genome of a virus into its particle is a major hurdle that any virus must overcome, HBV included, and understanding the mechanisms behind this process may prove valuable in understanding viral replication (1). This paper describes their process of uncovering how HBV packages its DNA genome into the mature viral particle (2).

Viral Particle Imaging

The paper by Gibes et al. 2026 describes their process of imaging Duck HBV (DHBV) and human HBV viral particles to uncover how the DNA inside these viruses is organized in a manner that allows them to fit inside such a small particle. DHBV was generally easy to use, which is why some of their experiments started using it rather than HBV (2). They did this by purifying DHBV and later HBV viral particles and imaging them using cryo-electron microscopy, which is used to take pictures of small particles such as viruses that would be too small for normal microscopes (Figure 1).

Figure 1: Cryo-em image of HBV particle. a. Fully developed HBV protein capsid. White shapes represent the geometric points of symmetry that pattern the capsid's outer layer. b. First layer of DNA (lavender) showing 11 concentric rings of DNA. c. Second layer of DNA (purple) showing 7 concentric rings of DNA. d. The third and final layer of DNA (blue) showing less-defined concentric rings of DNA (2).

Structural Analysis

They saw that the viruses contained their DNA in layers wrapped around protein, forming concentric DNA rings inside the viral particle (Figure 1 b-d). To generate this structure, the precursor RNA associates with the capsid, or viral shell, proteins. The RNA is then reverse transcribed into the DNA, which replaces the RNA. The structures they derived from their experiments hint that the process of copying the RNA into DNA may cause the rings of DNA to slide around the proteins, and as more layers are added, the DNA would become more rigid in its position in the viral particle (2). One of the ends of the capsid protein monomers that the DNA associates with is rich in arginine, a positively charged amino acid, which can interact with the negatively charged backbone of DNA to stabilize the negative charges that would end up being so tightly packed into the viral particle (Figure 2). 

Figure 2: Structures of charged molecules relevant to capsid stability a. Structure of L-arginine b. Structure of a portion of the DNA backbone, “Base” represents A, T, C, or G (2)

They speculate that the tightly packed DNA can also act as a spring inside the viral particle, so that when it infects a new liver cell, the force from the tightly wound DNA can cause the particle to burst open and facilitate genome entry into the cell (2). 

Conclusions

To better understand how HBV packages its entire genome into its viral particle, the authors imaged mature HBV and DHBV particles to visualize the organization of the DNA inside. They revealed that the DNA of HBV and DHBV is contained in layers of circular concentric loops. The authors speculate that the process of packaging the particle takes place during reverse transcription of the RNA segments that HBV creates following infection, and that the positively charged capsid proteins facilitate stabilizing the DNA inside the virus (2). 

Works Cited

1. Gibes N, Culhane K, Liu H, Xi J, Pionek K, Nair S, Loeb DD, Hu J, Zlotnick A, Wang JC. DNA synthesis inside the hepatitis B virus creates a high-energy spool. bioRxiv [Preprint]. 2026 Jan 23:2026.01.22.701187. doi: 10.64898/2026.01.22.701187. PMID: 41648622; PMCID: PMC12871785.

2. Watson AG, Mulay AS, Gill US. Chronic hepatitis B in 2025: diagnosis, treatment and future directions. Clin Med (Lond). 2025 Nov;25(6):100527. doi: 10.1016/j.clinme.2025.100527. Epub 2025 Nov 4. PMID: 41192690; PMCID: PMC12681808.