Investigating a Common Virus’ Damaging<br />Effect on Cells

By Tara Roberts

Dr. Lee FortunatoUniversity of Idaho biologist Lee Fortunato’s innovative new research method reveals how a virus responsible for birth defects co-opts a cell’s ability to repair itself.

Fortunato, an associate professor of biological sciences, has spent more than 15 years studying human cytomegalovirus (HCMV), and her recent research has been published in the Public Library of Science’s “Pathogens” journal. Though HCMV is common – 50-80 percent of humans are carriers – it can cause severe birth defects if a fetus is infected in utero.

Approximately 1 percent of babies born annually in the United States – about 40,000 – are infected with HCMV. Among them, some 4,000 are born with such conditions as loss of vision and hearing, cerebral palsy, mental retardation and microcephaly, or small head size. Another 4,000 develop problems, such as progressive hearing loss, during childhood.

Fortunato’s work examines the effects of the virus on cellular DNA repair mechanisms. It could lead to an antiviral therapy and new understanding of the ways infections caused by various disease-causing organisms defeat cells’ defenses.

When HCMV infects a cell, it sets up centers inside the nucleus to replicate itself. The proteins that repair cellular damage – including P53, known as the “guardian of the genome” – become sequestered inside these centers.

Fortunato hypothesized the virus was taking over the cell’s repair mechanisms and using them for itself, leaving the cells unable to fix themselves if they incurred damage or kill themselves to stop the spread of the virus.

Selective removal of CPDs (in green) from the viral DNA replication centers (BrdU label in red) after UV irradation of HCMV-infected cellsPhoto: Selective removal of DNA damage (in green) from the viral DNA replication centers (BrdU label in red) after UV irradation of HCMV-infected cells

She exposed infected cells to UV irradiation, which equally damaged the cellular and viral DNA. She then recorded the rate of repair using an established assay.

Researchers can use agarose gel assays to evaluate the amount of damage present in a DNA strand. In these assays DNA is cut into small fragments by a UV damage-specific enzyme. The smaller the fragments, the more damage to the DNA. Running the damaged, digested DNA on the gel results in a smear. As the UV damage is repaired, the smear disappears and is replaced by a single band (representing large, repaired fragments) at the top of the gel.

Unlike typical studies that don't differentiate between cellular and viral DNA in the infected cells, Fortunato's test examined repair in both genomes separately, yet simultaneously. The test showed the cellular machinery repaired the viral DNA far more quickly and efficiently than the cellular DNA.

“We’ve shown that, at least with UV irradiation of an infected cell, the viral genome is preferentially repaired, but the cellular genome is not repaired,” Fortunato says. “That could have ramifications for an infected fetus.”

Fortunato says it is very likely the virus keeps cells from repairing themselves when damaged by factors other than UV radiation. Other researchers could use her method for comparing DNA damage to prove it.

Following Fortunato’s method for testing cellular DNA separately from invasive DNA could open doors in other research as well. And, in the long term, Fortunato’s work could greatly reduce the negative ramifications of HCMV infections.

Fortunato’s research is supported by long-term funding from the National Institutes of Health and through larger NIH-supported grants designated for building Idaho’s biomedical research capacity.

Reducing the risk of HCMV infection

A woman who contracts human cytomegalovirus (HCMV) during pregnancy has a greater chance of passing the virus through the placenta to her unborn baby. The problem is compounded by the fact that a person who is infected by one strain of HCMV can contract another, and the virus can lie latent in the body after initial infection but can be reactivated later.

“The real kicker is, our immune system should be able to fight this virus off,” says Lee Fortunato, a U-Idaho biologist who studies the virus. “It clearly doesn’t do a very good job at this.”

But until an antiviral or vaccine is developed, Fortunato says, pregnant women can reduce their risk of contracting HCMV with simple measures. Since many women contract the virus from their older children, The Centers for Disease Control recommends that they avoid sharing utensils or kissing their children on the lips during pregnancy. The CDC also recommends frequent hand washing, especially after diaper changes.