ANN ARBOR— What evil lurks in the hearts of cells? Erle
Robertson and Murray Cotter know.These University of Michigan scientists have discovered how some viruses can hide inside the nucleus of human cells for long periods of time—without producing symptoms or triggering an immune response—by attaching to host cell chromosomes. The virus survives by going dormant until a weakened immune system allows infected cells to begin multiplying wildly again.

In an article to be published in the Nov. 25, 1999, issue
of Virology, Robertson and Cotter describe a series of
experiments with Kaposi’s sarcoma-associated herpesvirus or
KSHV—a human virus associated with a type of cancer
called Kaposi’s sarcoma. In these studies, U-M scientists
found a protein expressed by one gene on the virus that
builds a biochemical docking station linking viral DNA to
the chromosomes of lymphoma cells. The U-M study is the
first to identify a specific tethering mechanism between a
virus and its host cell.

KSHV, also known as human herpesvirus 8, is one of a family
of gammaherpesviruses known to remain dormant in humans
long after the initial infection is over. Other similar
viruses include the Epstein-Barr virus, the human papilloma
virus which causes cervical cancer, and viruses responsible
for hepatitis B and hepatitis C.

“We’ve always suspected that latent viral DNA couldn’t
survive long term within cells without some type of
tethering,” said Robertson, Ph.D., assistant professor of
microbiology and immunology in the U-M Medical School.
“But the latency mechanism for these viruses has been a
black box. Now we have a key that will get us in the front
door.”

Using cultures of lymphoma cells infected with KSHV, Cotter
and Robertson identified a protein called the latency-
associated nuclear antigen or LANA, which is expressed by
one of approximately 80 genes encoded by the virus. They
found that LANA binds to three regions of the KSHV genome,
but is most likely to lock onto one specific region for
tethering the virus to host chromosomes.

In addition to viral DNA, U-M scientists found that LANA
also binds to histones—small proteins that link bundles
of DNA called nucleosomes together to make chromatin
fibers, which are folded and packed to form chromosomes.

“The results suggest a biochemical mechanism which binds
elements of viral DNA to host chromosomes through the
interaction of LANA, histone H1 and possibly other
chromosomal proteins,” Robertson said.

Robertson has evidence for a similar tethering mechanism in
the Epstein-Barr virus, which infects immune system cells
called B-lymphocytes. Associated with several varieties of
cancer, including breast cancer, Epstein-Barr virus is
found in more than 90 percent of the world’s population.

In most people, a healthy immune system keeps the virus
suppressed. If something upsets the balance between virus
and immune response, however, the virus can re-activate.
The trigger that signals a dormant virus to begin
multiplying and infecting new cells remains unknown, according to Robertson.In previous studies with Epstein-Barr virus, scientistsidentified a protein called EBNA1, which binds to B-lymphocyte chromosomes. Since ENBA1 is expressed in allEBV-infected cells and LANA is expressed in all KSHV-infected cells, Robertson believes they may have similarfunctions. “We haven’t linked all the pieces yet, but itis extremely likely that EBNA1 is part of a similar tethering mechanism for Epstein-Barr viral DNA,” he said.

“If there are two viruses with the same mechanism, then
there probably are more.”

HIV, the virus that causes AIDS, is a different type of
virus and is unrelated to the U-M study, adds Robertson.
However, Kaposi’s sarcoma is a common cancer in people
whose immune system has been suppressed by the AIDS virus.

In future research, U-M scientists will try to find LANA’s
exact binding site on Kaposi’s viral DNA and on histone
proteins. “Once we understand the biochemistry of the
tethering site, we can start developing therapeutic agents
to block it. Blocking the binding site could mean the
difference between just treating symptoms and eradicating
these viruses from the host population,” added Murray A. Cotter II, a graduate student in theU-M Medical School.

Cotter also hopes to apply what he learns from KSHV biology
to gene therapy research. “The central question in gene
therapy is how do you stabilize foreign genes in a cell’s
nucleus, so they will express beneficial proteins over long
periods of time,” he said. “This tethering mechanism may
give us clues that we could use to develop more effective
gene therapy vectors.”

The U-M has applied for a patent on the viral tethering
mechanism. The research was supported by the National
Cancer Institute, the American Heart Association, the
Leukemia Society of America, and the U-M Health System’s
Comprehensive Cancer Center.