Paul Basilio, MDLinx | February 09, 2018
Scientists at the Morgridge Institute for Research in Madison, WI, have discovered a promising new target to fight a class of viruses responsible for hepatitis C, Zika, polio, dengue, and SARS. The results were published in the journal Science Advances.
Masaki Nishikiori, a researcher in the institute’s virology group, showed for the first time that viruses create pores inside parts of the cell that are normally walled off in genomic replication. This process of “punching through cellular walls” allows the virus to operate across different parts of the cell to activate and regulate its replication.
The virology group is led by Paul Ahlquist, PhD, Morgridge investigator and professor of oncology and molecular virology at the University of Wisconsin-Madison.
The findings could be important news in the quest to develop broad-spectrum antivirals. Hundreds of viruses threaten human health, but today the only way to combat them is by targeting each individual strain, rather than finding a common weakness.
The study looks at positive-strand RNA viruses, which make up one-third of all known viruses. The pore-creating mechanism could be common across many or most members of this family of viruses.
“One exciting aspect of these results is that pores of different kinds in membranes are very important for many biological processes, and there are established drugs that interfere with them,” said Mr. Nishikiori. “We now recognize that this virus—and based on conserved features, likely most viruses in this class—depend on similar types of pores to replicate. This is a target we know how to interfere with.”
Current pore-blocking drugs, also referred to as channel blockers, are used in treating high blood pressure, certain neurological or psychiatric disorders such as Alzheimer’s disease, and other maladies.
Mr. Nishikiori used biochemical and molecular genetic approaches to reveal the virus’s capacity to create and employ pores in a cell membrane. He used an advanced bromovirus model that allowed viral replication in yeast cells, which provided a highly controllable system to modify and assess both virus and host-cell contributions.
Viruses hijack normal cell structures and functions to spread throughout the body. This class of virus, for example, always anchors its genome replication process to the membrane surfaces of cell subcompartments or organelles (in this case, the endoplasmic reticulum). It had been understood that this process occurred solely on one side of the membrane, outside of the organelle.
The study revealed that the conventional view is incomplete. Mr. Nishikiori found that an enzyme called ERO1, which resides exclusively inside the organelle, on the opposite side of the membrane, is crucial to promoting the viral replication process. Reduce ERO1 and viral replication goes down, and vice versa, he explained.
However, there was a surprise. How could an enzyme that was walled off from the virus by a solid membrane barrier activate viral growth? This was their first clue that something must be bridging the membrane. When combined with other insights, the team discovered that a key viral protein builds a pore or pipeline across the membrane, enabling ERO1 to affect viral replication on the other side.
Mr. Nishikiori previously found that the proteins creating these pores become strongly linked together by a chemical bond, called a disulfide bond. However, these bonds can only be created in an oxidized environment. This explains at least one purpose behind the pores, he said. They allow delivery of oxidizing power into the normally nonoxidized cytoplasm to form the disulfide bonds.
“When the viral protein creates this pore, it allows oxidants generated by ERO1 to leach from the organelle interior into the cytoplasm and create a plume of oxidizing power,” Mr. Nishikiori added.
Dr. Ahlquist speculated that the virus may use these strong linkages to keep the viral genome replication apparatus intact. Drawing on other recent findings, he said viral replication complexes inside cells are under significant pressure and at risk of breaking off before replication is complete. The strong bonds might be similar to the wire cage holding the cork in place on a champagne bottle, he noted.
Basic research on the mechanisms of viral replication is essential to the larger quest to find broad-spectrum antivirals, one of the holy grails of virology, Dr. Ahlquist explained.
“When you apply an over-the-counter antibacterial cream to a child’s scraped knee, it works even though you don’t know exactly which bacteria you’re fighting,” he said. “We don’t have anything like that for viruses; most of our antiviral vaccines and drugs are virus-specific. We need new approaches that target broadly conserved viral features to simultaneously inhibit many viruses.”
In a separate study published in Scientific Reports, researchers at University of California San Diego School of Medicine announced that sofosbuvir effectively protected and rescued neural cells infected by the Zika virus, and blocked transmission of the virus to mouse fetuses.
“There has been a lot of work done in the past year or so to address the Zika health threat. Much of it has focused on developing a vaccine, with promising early results,” said senior author Alysson Muotri, PhD, professor in the UC San Diego School of Medicine departments of Pediatrics and Cellular and Molecular Medicine, director of the UC San Diego Stem Cell Program, and a member of the Sanford Consortium for Regenerative Medicine. “But there is also a great need to develop clinical strategies to treat Zika-infected individuals, including pregnant women for whom prevention of infection is no longer an option. They represent the greatest health crisis because a Zika infection during the first trimester confers the greatest risk of congenital microcephaly.”
In tests using human neural progenitor cells (NPCs) — self-renewing, multipotent cells that generate neurons and other brain cell types — the scientists found that exposure to sofosbuvir not only rescued dying NPCs infected with the Zika virus, but restored gene expression linked to their antiviral response.
In subsequent tests using an immunodeficient mouse model infected by Zika, intravenous injections of sofosbuvir significantly reduced viral loads in blood serum compared with a placebo group. Moreover, fetuses of Zika-infected pregnant mice did not show detectable Zika virus amplification in the sofosbuvir-treated group.
“This suggests that one, the drug was well-tolerated by the Zika-infected pregnant mice, and two, more importantly, that it was able to arrest Zika replication in vivo and stop transmission from mother to fetus,” said Dr. Muotri.
The researchers emphasized that their findings are preliminary, and much more work needs to be done.