viruses

Holding on: Bond LSC scientist discovers protein prevents release of HIV and other viruses from infected cells

Shan-Lu Liu and Minghua Li, HIV Research at the Bond Life Sciences Center

Shan-Lu Liu, Bond LSC scientist and associate professor in the MU School of Medicine’s Department of Molecular Microbiology and Immunology. Courtesy Justin Kelley, University of Missouri Health System.

Shan-Lu Liu initially thought it was a mistake when a simple experiment kept failing.

But that serendipitous accident led the Bond Life Sciences Center researcher to discover how a protein prevents mature HIV from leaving a cell.

Proceedings of the National Academy of Sciences published this research online Aug. 18.

“It’s a striking phenomena caused by this particular protein,” Liu said. “The HIV is already assembled inside the cell, ready for release, but this protein surprisingly tethers this virus from being released.”

The TIM – T-cell/transmembrane immunoglobulin and mucin – family of proteins hasn’t received much attention from HIV researchers, but recent research shows the protein family plays a critical role in viral infections.  From Ebola and Dengue to Hepatitis A and HIV, these proteins aid in the entry of viruses into host cells.

But its ability to stop the virus from leaving cells remained unknown until now. Liu’s lab stumbled onto this finding in November 2011 when trying to create stable cells for a different experiment. After two months of troubleshooting the HIV lentiviral vector – where genes responsible for creating TIM-1 proteins were inserted into a cell to create a stable cell line that expresses the protein – Liu was confident the vector’s failure was not only interesting but also important.

Shan-Lu Liu and Minghua Li, HIV Research at the Bond Life Sciences Center

Minghua Li, coauthor of the study and an MU Area of Pathobiology graduate student. Courtesy Justin Kelley, University of Missouri Health System.

The lab spent the next two years trying to figure out what was happening. Minghua Li, an MU Area of Pathobiology graduate student, carried out experiments that confirmed the protein’s power to inhibit HIV-1 release from cells, reducing normal viral infection. His experiments showed TIM proteins prevent normal deployment of HIV, created by an infected cell, into the body to propagate.

TIM proteins stand erect like topiary on the outside and inside surfaces of T-cells, epithelial cells and other cells. When a virus initially approaches a cell, the top of each TIM protein binds with fats – called phosphatidylserine (PS) – covering the virus surface. This allows a virus, such as Ebola virus and Dengue virus, to enter the cell, infect and replicate, building up a population inside.

But as the virus creates new copies of itself, the host cell’s machinery also incorporates TIM proteins into new viruses. That causes problems for HIV as it tries to leave the cell. Now these proteins cause the viruses to bind to each other, clumping together and attaching to the cell surface.

“We see this striking phenotype where the virus just accumulates on the cell surface,” said Liu, who is also an associate professor in the MU School of Medicine’s Department of Molecular Microbiology and Immunology. “We consider this an intrinsic property of cellular response to viral infection that holds the virus from release.”

This model shows the  interaction between TIMs and PS among the round HIV virions, as well as that between viral producer cells. This collectively leads to accumulation of HIV virions on the plasma membrane on the outside of the cell. Courtesy Mingua Li.

This model shows the interaction between TIMs and PS among the round HIV virions, as well as that between viral producer cells. This collectively leads to accumulation of HIV virions on the plasma membrane on the outside of the cell. Courtesy Minghua Li.

Further research is needed to determine overall benefit or detriment of this curious characteristic, but this discovery provides insight into the cell-virus interaction.

“This study shows that TIM proteins keep viral particles from being released by the infected cell and instead keep them tethered to the cell surface,” said Gordon Freeman, Ph.D., an associate professor of medicine with Harvard Medical School’s Dana-Farber Cancer Institute, who was not affiliated with the study. “This is true for several important enveloped viruses including HIV and Ebola. We may be able to use this insight to slow the production of these viruses.”

The National Institutes of Health and the University of Missouri partially supported this research. Additional collaborators include Eric Freed, PhD, senior investigator with the National Cancer Institute (NCI) HIV Drug Resistance Program; Sherimay Ablan, biologist with the NCI HIV Drug Resistance Program; Marc Johnson, PhD, Bond LSC researcher and associate professor in the MU Department of Molecular Microbiology and Immunology; Chunhui Miao and Matthew Fuller, graduate students in the MU Department of Molecular Microbiology and Immunology; Yi-Min Zheng, MD, MS, senior research specialist with the Christopher S. Bond Life Sciences Center at MU; Paul Rennert, PhD, founder and principal of SugarCone Biotech LLC in Holliston, Massachusetts; and Wendy Maury, PhD, professor of microbiology at the University of Iowa.

Read the full study on the PNAS website and browse the supplementary data for this work. See more news on this research from the MU School of Medicine.

Viruses as Vehicles: Finding what drives

Graduate students Yuleam Song and Dan Salamango inoculate a bacteria culture in Johnson's lab. The inoculation takes a small portion of a virus and multiplies the sample, allowing researchers to custom-make viruses.

Graduate students Yuleam Song and Dan Salamango inoculate a bacteria culture in Johnson’s lab. The inoculation takes a small portion of a virus and multiplies the sample, allowing researchers to custom-make viruses.

By Madison Knapp | Bond Life Sciences Center summer intern

Modern science has found a way to turn viruses —tiny, dangerous weapons responsible for runny noses, crippling stomach pains and worldwide epidemics such as AIDS— into a tool.

Gene therapy centers on the idea that scientists can hijack viruses and use them as vehicles to deliver DNA to organs in the body that are missing important genes, but the understanding of virus behavior is far from exhaustive.

Marc Johnson, researcher at the Christopher S. Bond Life Sciences Center and associate professor of molecular microbiology and immunology in the MU School of Medicine, has been building an understanding of viral navigation mechanisms which allow a virus to recognize the kind of cell it can infect.

Johnson’s research specifically explores the intricacies of the viral navigation system and could improve future direction of gene therapy, he said.

 

Marc Johnson (left) with a post doctoral student that works in his lab. The lab does important research on the basic function and mechanisms of viral navigation and transport.

Marc Johnson (left) with Dan Salamango, a graduate student that works in his lab. The lab does important research on the basic function and mechanisms of viral navigation and transport.

Turning a virus into a tool

Conceptualized in the 1970s, gene therapy was developed to treat patients for a variety of diseases, including Parkinson’s, leukemia and hemophilia (a genetic condition that stops blood from clotting).

To treat disease using gene therapy, a customized virus is prepared. A virus can be thought of as a missile with a navigation system and two other basic subunits: A capsule that holds the ammunition and the ammunition itself.

The viral genetic material can be thought of as the missile’s ammunition. When a cell is infected, this genetic material is deployed and incorporated into the cell’s DNA. The host cell then becomes a factory producing parts of the virus. Those parts assemble inside the cell to make a new virus, which then leaves the cell to infect another.

The capsule is made of structural protein that contains the genetic material, and the navigation system is a protein that allows the virus to recognize the kind of cell it can infect.

 

Viral navigation

Gene therapy uses viruses to solve many problems by utilizing a virus’ ability to integrate itself into a host cell’s DNA; to do this successfully, researchers need to provide a compatible navigation component.

In the body, viruses speed around as if on a busy highway. Each virus has a navigation system telling it which cells to infect. But sometimes if a virus picks up the wrong type of navigation system, it doesn’t know where to go at all.

“What you can do is find a virus that infects the liver already, steal its navigation protein and use that to assemble the virus you want to deliver the gene the liver needs,” Johnson said. “You can basically take the guidance system off of one and stick it onto another to custom design your virus.”

But this doesn’t always work because of incompatibility among certain viruses, he said.

Johnson and his lab are working to understand what makes switching out navigation proteins possible and why some viruses’ navigation systems are incompatible with other viruses.

“I’m trying to understand what makes it compatible so that hopefully down the road we can intelligently make others compatible,” Johnson said.

 

The right map, the right destination

Johnson creates custom viruses by introducing the three viral components—structural protein, genetic material, and navigation protein—to a cell culture. The structural protein and genetic material match, but the navigation component is the wild card. It could either take to the other parts to produce an infectious virus, or it could be incompatible.

Johnson uses a special fluorescent microscope to identify which viruses assembled correctly and which didn’t.

A successful pairing is like making a match. If a navigation protein is programmed to target liver cells, it’s considered a successful pairing when the virus arrives at the liver cell target location.

The scope of gene therapy continues to widen. Improved mechanisms for gene therapy, and greater knowledge of how a navigation protein drives a virus could help more people benefit from the vehicles viruses can become.

Johnson uses several high-profile model retroviruses, including human immunodeficiency virus (HIV), which affects an estimated 35 million people worldwide each year, according to the World Health Organization.

Understanding nuances of HIV in comparison to other viruses allows Johnson to pick out which behaviors might be common to all retroviruses and others behaviors that might be specific to each virus.

Johnson said his more general approach makes it easier to understand more complex viral features.

“If there are multiple mechanisms at work, it gets a little trickier,” Johnson said. “My angle is more generic, which makes it easier to tease them apart.”

Supervising editor is Paige Blankenbuehler