Computational Evolutionary Analysis of the Overlapped Surface (S) and Polymerase (P) Region in Hepatitis B Virus Indicates the Spacer Domain in P Is Crucial for Survival

Ping Chen,Wei Fang,Yun Gan,Jiafu Li,Na Han,Simon Rayner,Fei Zhao,Kanghong Hu,Yury E Khudyakov

doi:10.1371/journal.pone.0060098

Abstract

IntroductionThe Hepatitis B Virus (HBV) genome contains four ORFs, S (surface), P (polymerase), C (core) and X. S is completely overlapped by P and as a consequence the overlapping region is subject to distinctive evolutionary constraints compared to the remainder of the genome. Specifically, a non-synonymous substitution in one coding frame may produce a synonymous substitution in the alternative frame, suggesting a possible conflict between requirements for diversifying and purifying forces. To examine how these contrasting requirements are balanced within this region, we investigated the relationship amongst positive selection sites, conserved regions, epitopes and elements of protein structure to consider how HBV balances the contrasting evolutionary pressures.Methodology/Results323 HBV genotype D genome sequences were collected and analyzed to identify sites under positive selection and highly conserved regions. Epitopes sequences were retrieved from previously published experimental studies stored in the Immune Epitope Database. Predicted secondary structures were used to investigate the association between structure and conservation. Entropy was used as a measure of conservation and bivariate logistic regression was used to investigate the relationship between positive selection/conserved sites and epitope/secondary structure regions. Our results indicate: (i) conservation in S is primarily dictated by α-helix elements in the protein structure, (ii) variable residues are mainly located in PreS, the major hydrophilic region (MHR) and the C-terminus, (iii) epitopes in S, which are directly targeted by the host immune system, are significantly associated with sites under positive selection.ConclusionsThe highly variable spacer domain in P, which corresponds to PreS in S, appears to act as a harbor for the accumulation of mutations that can provide flexibility for conformational changes and responding to immune pressure.

Highlights

The Hepatitis B Virus (HBV) genome contains four open reading frames (ORF), S, P, C and X
Identification of Positive Selection in P and S The six site models implemented in the PAML software package were compared (M3 to M0, M2 to M1, and M8 to M7) by the likelihood ratio test
For P, 86.7 percent (13 out of 15) of the positive selection sites are located in the spacer domain, while the reverse transcriptase (RT) domain only contains two sites (13.3%)

Summary

Introduction

The Hepatitis B Virus (HBV) genome contains four ORFs, S (surface), P (polymerase), C (core) and X. A non-synonymous substitution in one coding frame may produce a synonymous substitution in the alternative frame, suggesting a possible conflict between requirements for diversifying and purifying forces To examine how these contrasting requirements are balanced within this region, we investigated the relationship amongst positive selection sites, conserved regions, epitopes and elements of protein structure to consider how HBV balances the contrasting evolutionary pressures. Gene overlapping is a common strategy adopted by many viruses to reduce their genome size and maximize their encoding capacity This inevitably constrains the independent evolution of the individual reading frames as a mutation with little effect on one gene may cause severe or even fatal changes on the cognate overlapping gene. In an overlapping region, if one gene undergoes adaptive evolution (positive selection) with a high ratio of nonsynonymous nucleotide mutations (dn/ds .1), the cognate gene often undergoes purifying selection (negative selection; dn/ds ,1)

Methods

Results

Conclusion