Molecular biology of the hepatitis B virus for clinicians

Abstract

Hepatitis B virus (HBV) infection is one of the major global health problems, especially in economically under-developed or developing countries. HBV infection can lead to a number of clinical outcomes including chronic infection, cirrhosis and liver cancer. It ranks among the top 10 causes of death, being responsible for around 1 million deaths every year. Despite the availability of a highly efficient vaccine and potent antiviral agents, HBV infection still remains a significant clinical problem, particularly in those high endemicity areas where vaccination of large populations has not been possible due to economic reasons. Although HBV is among the smallest viruses in terms of virion and genome size, it has numerous unique features that make it completely distinct from other DNA viruses. It has a partially double stranded DNA with highly complex genome organization, life cycle and natural history. Remarkably distinct from other DNA viruses, it uses an RNA intermediate called pregenomic RNA (pgRNA) and reverse transcriptase for its genome replication. Genome replication is accomplished by a complex mechanism of primer shifting facilitated by direct repeat sequences encoded in the genome. Further, the genome has evolved in such a manner that every single nucleotide of the genome is used for either coding viral proteins or used as regulatory regions or both. Moreover, it utilizes internal in-frame translation initiation codons, as well as different reading frames from the same RNA to generate different proteins with diverse functions. HBV also shows considerable genetic variability which has been related with clinical outcomes, replication potential, therapeutic response etc. This review aims at reviewing fundamental events of the viral life cycle including viral replication, transcription and translation, from the molecular standpoint, as well as, highlights the clinical relevance of genetic variability of HBV.

Keywords: AUG, translation start codon; BCP, basal core promoter; CHB, chronic hepatitis B infection; DR, direct repeat; EBP, enhancer binding protein; EN, enhancer; ER, endoplasmic reticulum; HBV, hepatitis B virus; HBsAg; HCC, hepatocellular cancer; Hepadnavirus; IL, interleukin; LEF, liver enriched factors; LHB, large envelope protein; MHBs, middle hepatitis B surface antigen; MHR, major hydrophilic region; ORF, open reading frames; PC, precore; RT, reverse transcriptase; SHBs, small hepatitis B surface antigen; TGF-α, transforming growth factor-α; TNF-α, tumor necrosis factor-α; TP, terminal protein; WHV, woodchuck hepatitis virus; cccDNA, covalently closed circular; dGMP, deoxyguanosine monophosphate; genotype; pHSA, poly-human serum albumin; pgRNA; pgRNA, pregenomic RNA; rcDNA; rcDNA, relaxed circular DNA.

Figure 1

Schematic representation of different forms of infectious and non-infectious hepatitis B virus particles. (a) Complete viral particle, (the 42–47 nm Dane particle), (b & c) two species (filamentous and spherical respectively) of non-infectious 20 nm surface antigen particles. ‘Pol’ indicates HBV polymerase protein. Figure not according to the scale.

Figure 2

Schematic diagram of the genome and translational map of HBV. The innermost red circles depict the rcDNA with the reverse transcriptase/polymerase (Pol) attached to the 5′ end of the complete minus strand DNA (solid red circle) and a capped RNA oligomer (wavy red line) attached to the 5′ end of the incomplete plus strand DNA (dotted red circle). The positions of the direct repeats (DR1 and DR2) and enhancers (EN1 and EN2) are also indicated. The green line indicates the viral genome positions in nucleotides (approximate). The four protein-coding regions are shown between the green and red circles by colored semi-circular arrows. They include the precore (PC) and core genes (violet), the polymerase gene (blue), the X gene (aqua) and the envelope genes preS1, preS2, and S (orange). Genome positions may change depending upon the genotype of the HBV genome (modified from Nassal 2008). The outermost semi-circular lines with arrowheads represent the 4 RNAs (genomic and subgenomic) corresponding to the ORFs. The arrowheads indicate the positions of different initiation codons within each ORF.

Figure 3

Schematic diagram showing life cycle of the HBV.

Figure 4

Consecutive steps of HBV genome replication. (a) Translation of pgRNA and initiation of minus strand reverse transcription using the TP domain of Pol and bulge region of ‘ε’ as template. (b) First primer shift-translocation of nascent minus strand to 3′ end of pgRNA and base pairing with DR2. (c) Completion of minus strand using pgRNA as template and simultaneous degradation of pgRNA. (d) Second primer shift and template shift—reallocation of RNA primer to the 5′ end of the minus strand (DR2), followed by circularization of the genome due to redundant sequences present in the minus strand DNA. (e) Formation of relaxed circular DNA (rcDNA)-incomplete elongation of the plus strand DNA using minus strand DNA as template. (f) Infection and repair of rcDNA to covalently closed circular DNA.

Figure 5

Schematic diagram showing the stem loop structure of the encapsidation sequence of the HBV precore genome. The relation between nucleotides at position 1858 and 1896 is shown. Positions having variable nucleotides within the structure are indicated.

Figure 6

SchematicIm diagram showing the functional domains of the HBV polymerase (Pol) protein, and the active domains within the polymerase/reverse transcriptase (RT) domain. ‘TP’ indicate terminal protein. Position of different antiviral resistance mutations within the polymerase/RT domain is also shown below [adapted and modified from Locarnini and Yuen, 2010].