MappingofB-Cell Epitopes oftheHumanHepatitis B VirusX Protein

TheimmuneresponsetotheX protein ofhumanhepatitis B virus (HBV)was studied byepitope mapping byusing asetofMS2-HBxfusion proteins andsynthetic peptides. Antibodies inseraofpatients withacuteand chronic HBV infection showed amultispecific immuneresponse.Eachserum contained antibodies toadifferent setofepitopes, whichtaken together covermostoftheHBxsequence.Someoftheepitopes weredetectable only byimmunoblotting withfusion proteins; others were detectable onlybyan enzyme-linked immunosorbent assay(ELISA) withsynthetic peptides. Thecarboxy-terminal halfoftheHBx protein was preferentially recognized byantibodies frompatients withchronic hepatitis andcontained ashort immunodominant antigenic region withatleast twomajornonoverlapping epitopes. Anti-HBx antibody titers as revealed bypeptide ELISAswere highest andmostfrequent inpatients withchronic hepatitis andusually lowinacutely infected patients andasymptomatic carriers. Thedatademonstrate a remarkable qualitative andquantitative heterogeneity ofthehumoral HBximmuneresponsewhich canbemonitored byHBx-specific peptide ELISAs. Suchtests may becomeuseful diagnostic tools. Hepatitis B viruses (HBVs)havebeenfoundinhumans andseveral animalspecies suchas woodchucks, ground squirrels, Pekinducks, andgreyherons (14,41,45).Infectionwiththeseviruses oftenleadstoacuteor chronic infection andvarious formsofliver diseases, butitmay also leadtoan asymptomatic chronic carrier state(for a review, seereference 7).Moreover, chronic infection with mammalian, butnotavian, HBVsisassociated with a highrisk for development ofhepatocellular carcinoma (HCC)(1,41). HBVs arenotcytopathogenic, andtherefore itisgenerally assumedthatviral hepatitis ismediated primarily byimmune defense mechanisms. Theimmuneresponsehasbeenstudiedinthemostdetail forvirus componentsexpressed in large amounts.B-andT-cell epitopes havebeenidentified fortwoantigens encodedbytheviral C gene,themajor nucleocapsid protein (HBcAg), andthee antigen (HBeAg) (27, 28,39), a protein whichissecreted intotheserum and whichisnotassociated withviral particles (17). Circumstantialevidence suggests an important roleoftheimmune responsetothese antigens intermsofvirus elimination and pathogenicity, andantibodies tobothantigens are early diagnostic markers ofHBV infection (27, 28). Similar studies havebeenperformed on theimmune responsetothethree envelope proteins, pre-Sl, pre-S2, and HBsAg.B-andT-cell epitopes havebeenmapped, andthe participation ofantibodies andT cells toenvelope epitopes inimmune-mediated defense mechanisms andtheuse of envelope proteins asvaccines havebeenextensively investigated (27, 28). Theroleoftheimmuneresponse tominorviral proteins, suchas theviral polymerase andgenome-linked protein