Chronic hepatitis B virus (HBV) infection affects around 400 million people worldwide and greatly increases risk of hepatocellular carcinoma (HCC) 
. The molecular mechanism through which chronic HBV infection contributes to hepatocarcinogenesis remains incompletely understood. Transgenic mice expressing HBV large surface protein that suffer from chronic hepatitis due to different mechanisms eventually develop HCC, suggesting that HBV may be carcinogenic merely by virtue of being an inciter of immune-mediated inflammation and hepatocyte damage 
. Chronic liver necroinflammation is common in mouse models of spontaneous HCC from targeted deletion of various genes 
, which may or may not closely associated with genetic aberration in human HCC. Besides chronic liver inflammation, HCC is also known to occur in HBV-infected people in the absence of prolonged severe liver injury or cirrhosis 
; among all people with chronic hepatitis, those with HBV infection had a 7-fold greater risk for HCC than those without HBV infection 
. Thus although severe hepatitis, of whatever cause, can cause HCC, in natural infections, HBV may play a more direct role in hepatocarcinogenesis. Transgenic mice overexpressing X protein from an HBV subgenomic fragment have been showed to develop HCC 
, but a number of transgenic lines expressing X protein from several constructs do not develop HCC 
. Importantly, transgenic mice expressing the entire HBV genome have not been shown to increase the incidence of HCC 
, unless the mice are treated with carcinogens 
. Thus it is unclear if HBV is directly carcinogenic by itself.
The surface proteins constitute the envelope proteins of HBV. Three forms, differing in the initiating codons, are synthesized from a single open-reading frame, with the large form comprising the preS1+preS2+S regions, the middle comprising the preS2+S regions, and the small form having only the S region (). The preS1 promoter-driven transcripts code for the large surface protein, while the S promoter-driven transcripts code for the middle and small surface proteins. The large and small forms are essential for viral morphogenesis and infectivity, but the middle form is dispensable 
. PreS2 mutants contain in-frame deletions in the N-terminus of the preS2 region as well as a frequent missense mutation of the preS2 start codon. These two mutations result in loss of expression of the middle surface protein and the synthesis of an internally deleted large surface protein and are likely selected because of immune pressure 
. PreS2 mutants also produce a moderately decreased amount of the small surface protein because the deletion removes a small downstream region of the S promoter 
(). Recent clinical data have shown that preS2 mutants are found at much higher frequencies in people with HCC than those without 
, and that these mutants are present in apparently clonal nodules of hepatocytes in infected livers 
. To test if these mutants play a causal role in the genesis of HCC, we generated transgenic mice containing such a mutant HBV genome.
Expression and replication of Mutant 1 HBV in transgenic mice.