Formation of semi‐enveloped particles as a unique feature of a hepatitis B virus PreS1 deletion mutant

Abstract

Summary Background Naturally occurring variants with deletions or mutations in the C‐terminal PreS1 domain from hepatitis B virus (HBV) chronically infected patients have been shown to promote HBsAg retention, inhibit HBsAg secretion and change the extracellular appearance of PreS1‐containing HBV particles (filaments and virions). Aims To study the impact of N‐terminal deletion in preS1 domain on viral secretion and morphogenesis. Methods An HBV mutant with 15 amino acids (aa 25‐39) deletion in N‐terminal preS1 was isolated. Intracellular and extracellular HBsAg were quantified by Western blot. Subcellular HBsAg distribution was analysed by confocal laser scanning microscopy. The viral morphology was characterised by sucrose density gradient ultracentrifugation, Western blot, electron microscopy, HBV mixed ELISA and HBV particle gel essay. Results Expression of this mutant genome released higher amounts of HBsAg in the form of shorter filaments. A significant fraction of semi‐enveloped virions was observed in the supernatant that has been unprecedented so far. Stepwise insertion of aa 25‐31, aa 32‐39 and aa 25‐39 increased the length of filaments. The rescue of aa 25‐31 and aa 25‐39 drastically reduced the amounts of extracellular HBsAg and semi‐enveloped virions, while such effects could not be observed after insertion of aa 32‐39, arguing against a simple spacer function of this region. The deletion and rescued mutants do not differ in subcellular HBsAg distribution and colocalisation with ER, Golgi and multivesicular bodies markers arguing against differences in release pathways. Conclusion N‐terminal PreS1‐domain (aa 25‐31) determines HBsAg secretion and triggers proper assembly of PreS1‐containing particles.

Intracellular Trafficking of HBV Particles

“…Based on a variant isolated from a patient suffering from chronic HBV that lacks aa 25-39 in the N-terminus of PreS1, it was found that the N-terminal PreS1 domain determines LHBs secretion and triggers proper assembly of HBV virions. This deletion caused a significant fraction of semi-enveloped viral particles, a variant that has not been described before [108]. The discrepancy between these studies could be based on the different experimental strategies.…”

“…The discrepancy between these studies could be based on the different experimental strategies. In the study from Jiang et al [108], HBV particle gel essay, HBV mixed ELISA and electron microscopy were mainly used to discriminate partially-enveloped viral particles from fully assembled viral particles. The above mentioned study by Bruss et al [106] performed immuno-precipitation and subsequent radioactive endogenous polymerase reaction (EPR) to quantify HBV viral particles, which fails to distinguish between partially-enveloped virions and intact enveloped virions.…”

“…The cryo-EM study of the tubular particles showed that the three proteins assemble as a mixed population of homo-and heterodimers, with spike-like features projecting from the membrane [6]. Deletion of aa 25-39 in the PreS1 domain of LHBs has been shown to be associated with a decreased assembly capacity, as reflected by the reduced length of filaments and appearance of incompletely enveloped viral particles [108]. Unlike SHBs, selective overexpression of the LHBs leads to the accumulation of LHBs and the inefficient secretion of HBsAg in the supernatant.…”

Editorial: HBV—the naked truth? Authors’ reply

“…The authors' declarations of personal and financial interests are unchanged from those in the original article …”

Editorial: HBV—the naked truth?

“…In a recent issue of AP&T, Jiang et al described the characterisation of an envelope deletion variant of hepatitis B virus (HBV) isolated from a patient. The authors showed that the deleted amino acid domain 25‐39 of PreS1 contributes to viral morphogenesis and affects hepatitis B surface antigen (HBsAg) secretion.…”