Quick Summary

• Systems biology addresses biological systems as a whole, rather than breaking them into smaller component parts
• Using it, researchers have gained new insights into how Dengue and Zika viruses attack their hosts
• They discovered a way to stop viruses from replicating in both human and mosquito cells, without harming the host cell

A new, holistic approach to biology is giving researchers new insights into how the Dengue and Zika viruses attack their hosts and, in the case of Zika, affect brain development. Published Dec. 13 in the journal Cell, the work may open up new ways to think about treating virus infections or mitigating their effects.

Systems biology addresses biological systems as a whole, rather than breaking them into smaller component parts. The approach encompasses high-throughput techniques in DNA sequencing and protein analysis (genomics and proteomics) as well as mathematical and computer modeling.

First author of the new study is Priya Shah, now an assistant professor in the Departments of Microbiology and Molecular Genetics and of Chemical Engineering at UC Davis. Shah carried out the work mostly as a postdoctoral researcher in Nevan Krogan’s laboratory at UC San Francisco.

Viruses have to accomplish three things, Shah said: “Get into a cell, replicate themselves, get out.” Dengue and Zika viruses belong to the same family, the flaviviruses, which are spread by mosquitoes.

“The fact that the viruses can accomplish these tasks in two different hosts separated by hundreds of millions of years of evolution is pretty interesting,” Shah said.

Both viruses have just ten protein-coding sequences in their genomes – so they work by hijacking host proteins.

Tagging virus proteins

Shah and colleagues tagged virus genes with a marker, then introduced them into human and insect cells where they produced proteins. Then they extracted broke up the cells and used the marker to purify virus proteins, along with the host proteins to which they stuck. After analyzing this mix of proteins by mass spectrometry, Shah could build a map of which human or insect proteins were interacting with virus proteins.

A number of interesting findings fell out of the analysis, she said.

Both dengue and Zika viruses have a protein called NS (“nonstructural”) 5, a polymerase responsible for copying the virus genome. NS5 is known to move between the host cell cytoplasm and the nucleus, although both viruses copy their genomes in the cytoplasm and apparently don’t need to enter the nucleus.

Shah and colleagues found that NS5 interacts with a protein in the human nucleus called PAF1C, which regulates immune response genes in the cell. NS5 blocks PAF1C’s immune role in the immune response and allows the viruses to reproduce more rapidly.

Mutant flies with small brains

Previously little studied, Zika virus came to wide attention in 2015 when it was linked to severe birth defects, especially microcephaly, in infants in Brazil.

Shah found that a Zika protein called NS4A interacts with a protein called ANKLE2, which has been linked to microcephaly.

ANKLE2 was discovered by Hugo Bellen’s laboratory at Baylor College of Medicine in Houston as a mutant that causes small brain size in Drosophila flies. When Bellen’s team compared the fly ANKLE2 to data from patients with unexplained defects in brain development, they found some cases of hereditary microcephaly linked to the human version of ANKLE2. And the cases look remarkably similar to the small heads of Zika-induced microcephaly.

When Bellen’s group tested Shah’s Zika NS4A genes in flies they found that they got the same “small-brain” phenotype as in flies with mutated ANKLE2.

“It suggests that the ANKLE2/NS4A interaction is important to developing microcephaly,” Shah said. “It’s probably one of several mechanisms that contribute to birth defects in Zika virus infection.”

NS4A from both dengue and Zika viruses also interacts with a protein complex called SEC61, which is involved in moving proteins around in the cell and especially putting proteins into membranes. SEC61 is found in both human and mosquito cells – so this interaction is important for each virus in both of their hosts.

Shah and colleagues found that they could use an inhibitor to selectively cut off SEC61 function and stop viruses from replicating in both human and mosquito cells, without harming the host cell.

This could be a route to therapy, Shah said, by making it harder for the virus to multiply. Because it targets a host process vital to the virus, rather than the virus itself, it should be harder for the virus to develop resistance to such a drug.

At UC Davis, Shah plans to continue working on the Zika/ANKLE2 problem, both in flies and in new animal models, such as zebrafish. As vertebrates with a more developed brain and spinal column, zebrafish are evolutionarily closer to us – and they are also transparent for the first few days of life, making it easier to watch how their brains develop.

The researchers also plan to continue work on NS5 and PAF1C, including if other flaviruses can also manipulate the immune response in this way.

Additional authors on the paper are: Nichole Link, Baylor College of Medicine; Gwendolyn Jang, Phillip Sharp, Danielle Swaney, Jeffrey Johnson, John Von Dollen, Laura Satkamp, Billy Newton, Ruth Hüttenhain, Orly Laufman, Michel Tassetto, Michael Shales, Erica Stevenson, Leila Shokat, A. Jeremy Willsey, Katherine Pollard, Jack Taunton, Raul Andino, Tierney Baum, and Amanda Everitt at UCSF and the J. David  Gladstone Institutes, San Francisco; Tongtong Zhu, Sebastian Aguirre, Shashank Tripathi, Laurence Webb, Ivan Marazzi, Ana Fernandez-Sesma, Icahn School of Medicine at Mount Sinai, New York; Holly Ramage and Sara Cherry, University of Pennsylvania; Marine Petit, UC Davis; Gabriel Iglesias and Andrea Gamarnik, Fundación Instituto Leloir-CONICET, Buenos Aires, Argentina; and Vinod Balasubramaniam, Monash University Malaysia.

The work was supported by grants from the NIH.