The Molecular Basis of Viral Replication 
and Pathogenesis 
Donald E. Ganem, M.D. — Associate Investigator 
Dr. Ganem is also Professor of Microbiology and Immunology and of Medicine at the University of 
California, San Francisco. He received an A.B. degree in biochemistry from Harvard College and the M.D. 
degree from Harvard Medical School. Following his clinical training in infectious diseases, he did 
postdoctoral research training in the laboratory of Harold Varmus at UCSF. 
OUR laboratory studies the molecular mecha- 
nisms by which pathogenic human viruses 
infect the host and cause disease. We are espe- 
cially interested in those viruses that produce 
persistent infections and engender chronic pa- 
thology. Several fundamental questions underlie 
our work: How are persistent infections estab- 
lished and maintained? What factors regulate 
viral replication and spread? How does the persis- 
tent presence of virus evoke disease? 
Most of our work centers around the human 
hepatitis B virus (HBV) and its animal homo- 
logues. HBV is a small DNA virus that replicates 
principally in liver cells (hepatocytes) and pro- 
duces acute and chronic type B hepatitis. Most 
initial infections are transient: following a rela- 
tively brief period of liver injury (hepatitis), the 
immune system eliminates virus from the liver 
and invokes lasting immunity. However, 5-10 
percent of infections are not successfully elimi- 
nated. In these cases, viral replication persists in 
the liver for the life of the host, evoking various 
degrees of chronic liver injury that can lead to 
premature death from liver failure. Most strik- 
ingly, persistence of viral replication for several 
decades enormously increases the risk of liver 
cancer. 
There are many reasons to be interested in this 
remarkable infection. First is its great public 
health significance. Worldwide, there are over 
250 million chronic HBV carriers. In large re- 
gions of Asia and Africa, 10-15 percent of all hu- 
man inhabitants are persistently infected by the 
virus. Second, the virus replication cycle is dis- 
tinctly unusual: replication of the viral DNA ge- 
nome is accomplished via reverse transcription of 
an RNA intermediate. The HBV life cycle is thus a 
permuted version of the retroviral life cycle, and 
a fuller understanding of its details allows in- 
structive comparisons to be made with cognate 
steps in retroviral replication. More importantly, 
unraveling the mechanism of HBV replication 
should identify new potential targets for antiviral 
therapy. Finally, the nature of the link between 
chronic HBV replication and hepatocellular carci- 
noma represents one of the great unsolved prob- 
lems in human cancer biology, and a deeper un- 
derstanding of HBV persistence is expected to 
provide new clues to its solution. 
Our studies of HBV replication have examined 
many different steps in the viral life cycle. The 
initial step in all viral infections is binding of the 
virus to the cell surface receptor and its entry into 
the cell. Little is known about this step in HBV 
infection, but we have begun to explore it in a 
convenient animal model, the duck hepatitis B 
virus (DHBV) . By using recombinant DHBV sur- 
face proteins to look for cellular proteins that 
will bind them, we have identified a single host 
glycoprotein that will interact with DHBV parti- 
cles with high affinity and specificity. Such mole- 
cules are good candidates for the receptor, and 
we are actively attempting to clone the gene for 
this host protein to allow its more-detailed 
characterization. 
Next, internalized virus particles must be deliv- 
ered to the nucleus, where their DNA genome is 
transcribed. This is perhaps the most poorly un- 
derstood of all reactions in virology. For no virus 
has the cellular machinery involved in this essen- 
tial step been identified. Recently, we discovered 
that drugs affecting the integrity of cellular mi- 
crotubules block this step, implying that these 
structures are involved in the intracellular trans- 
port of subviral particles. This raises the exciting 
possibility that such particles might also be used 
to identify host proteins that recognize them and 
perhaps bind not only virus particles but normal 
cellular components (e.g., organelles) to cyto- 
skeletal motors for transport within the cell. 
Once in the nucleus, viral genes are tran- 
scribed into RNA to be transferred to the cyto- 
plasm for translation. Following translation, prog- 
eny particles assemble in the cytoplasm. In this 
remarkable reaction, viral RNA is packaged into 
subviral particles along with the viral reverse 
transcriptase. Surprisingly, this RNA packaging 
process requires the participation of the reverse 
transcriptase itself. To understand this key step 
better, we are currently attempting to reproduce 
this RNA recognition in vitro, using recombinant 
reverse transcriptase and cloned viral RNA. Sim- 
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