Thwarting the next viral onslaught using electron microscopy
In 1981 a UCLA doctor described cases of patients dying of strange fever, lung infection and pneumonia. Reports around the country carried headlines of unexplained fatal illnesses caused by severe damage to the immune system.
As more and more reports appeared both nationally and internationally, it seemed like anyone could be affected. Panic spread among the world population. People died by the thousands.
By 1985, every major region in the world had reported at least one case of a disease that became knows as Acquired Immune Deficiency Syndrome, AIDS. The cause was identified as a tiny but insidious pathogen that we now call the Human Immunodeficiency Virus, HIV.
There’s an ancient saying by the Chinese philosopher Sun Tzu who, 2,500 years ago said, “If you don’t know your enemy, and you don’t know yourself, you will succumb in every battle.” For somebody who was diagnosed with HIV in the 80’s and early 90’s it was an almost certain death sentence. We had little idea of how the virus worked or where it attacked. We just knew that it was deadly.
This is the virus. That is a human immune cell. It doesn’t look like the cell should lose this battle, does it? The virus is clever. It’s learned to penetrate the immune cell, rapidly spread from one cell to another, and ultimately cripple the immune system leading to AIDS.
How does such a tiny organism cause so much damage? The virus host arms race has fascinated scientists for over a century. It is both for the arsenal of weaponry that each side employs, and for the grave effects to human health.
The struggle plays out in a world that we don’t often think about. It’s the molecular world. The living world that we see around us that are flora and fauna is just the tip of the biological iceberg. Our macroscopic world is actually composed of microscopic cells thousands of times smaller.
Inside each cell, there’s yet another world orders of magnitudes smaller still. It’s the world of molecules. A molecule is simply a collection of atoms held together by strong chemical bonds.
Carbon dioxide is a molecule. It’s a molecule that you’re probably familiar with. Sucrose, table sugar, is a molecule. Molecules can be a million times smaller than a grain of sand.
My own path to studying molecules professionally developed in a somewhat circuitous manner. After several iterations of trying to select a direction in school, including a brief stint into the world of philosophy where I studied everything from ethics, logic and probably even the meaning of the word molecule, I decided to try my hands at chemistry.
Chemistry aims to understand molecular composition and reactivity. It was fascinating because I learned how molecules can be manipulated using standard chemical principles. It was really the hands-on experience in a lab where I learned how to put together and break up these seemingly invisible molecules, that I became really intrigued.
Around that same time, I started learning about proteins. These are much larger and more elaborate molecular compositions made up of thousands of atoms which perform most of the biological functions in a cell. I also learned about genes. These are the basic molecular units and basic blueprints of heredity, or DNA. Although, it can also be RNA.
I learned about viruses. They are essentially compositions of proteins and genes. Viruses are clever. At least from our human perspective they can be quite destructive.
The fact that such clever organisms can be put together from these really basic protein and genetic building blocks, these basic constituents, was when I realized how much intricacy there was in the molecular world. I was hooked.
My lab at the Salk Institute studied the molecular world of proteins associated with HIV. Like any virus, HIV is a composition of proteins and genes. It’s infamous for the amount of devastation that it’s caused.
By the end of 2018, there would have been about 40 million people who would succumb to the disease. About the same number would be infected with the virus.
The virus has left behind a deadly mark. There are countries today with 25% infectivity rates. In the molecular world of HIV there’s a struggle that plays out between the virus and its host, the human immune cell.
The virus contains an arsenal of weaponry. The molecular arsenal in the molecules are viral proteins. Once the virus penetrates the cell, a group of proteins will then transport the viral genetic information, its DNA, blueprints for the virus’s own mass production, into the heart of the cell, the nucleus.
Once there, another group of proteins will then hijack the host DNA, and through a carefully coordinated set of chemical reactions will encode or integrate viral DNA with that of the host. We refer to this process as viral integration.
The virus becomes a part of you. Integration is one reason for the virus’s success and our failure to thwart it. In our effort to fight HIV, a key step or process that we need to understand is how integration works at the molecular level. How do we do this?
You’ve all heard the saying, “A picture is worth 1,000 words.” The same can be said of the molecular world. If we want to understand how these molecular machines work, we need a technology that allows us to observe them.
To gain glimpses into this tiny world, my lab uses a tool called an electron microscope. It’s simply a powerful microscope that uses electrons instead of light to image proteins with up to a million-fold magnification.
We can’t quite yet see these proteins in a cell, mostly because of the complexity of the environment. I’ll actually get to that a bit later. We can make the proteins in a lab. We can isolate them, purify them and visualize them in the microscope. Through image processing operations, we can come up with their atomic structure in three-dimensional representation.
Through our research, but as always building on the work of many others in the field, we’ve described how a group of HIV proteins mediates the integration of viral DNA with that of the host. We’re beginning to understand how this process works at the molecular level. How the virus occupies the cell and establishes its own molecular factories behind its enemy lines.
What did this process teach us? What did this knowledge teach us? The atomic level description of the viral protein arsenal that we obtained using an electron microscope provides us with a blueprint for understanding how to design and improve therapeutics that block protein function.
Modern medicines are simply small chemical molecules that target proteins in their Achilles heel, blocking their ability to perform harmful chemical reactions. Many medicines have been developed to fight HIV. The problem is, they’re designed to merely suppress the amount of circulating virus.
For 33 years after the development of the first therapeutic, there remains no cure. Sun Tzu also said, “If you know yourself but not the enemy,” and let me just add know the enemy but not yourself, “then for every victory gained, you will also suffer a defeat.”
We need to learn more. Prior to electron microscopy, determining structures mediating viral integration was extremely challenging. Now we’re doing this routinely. Moreover, we’re focusing not only on understanding their structure, but their dynamics and their interactions with host proteins. So, we’re zooming in on the actual battlefield. We’re using this information to design the next generation of therapeutics.
The state of the art is also about how much one can see. Recently, my lab described viral protein structures at one of the highest recorded resolutions using an electron microscope for biological samples.
We’re literally beginning to distinguish individual atoms like carbon, nitrogen, oxygen and sulfur. When we saw this data for the first time, I was in disbelief. I would have never thought that you could see such high level of detail for biological samples inside of an electron microscope. This is critical because, the more we see, the better our therapeutic blueprints.
We’re behind. While the technology I’ve described to you is ground breaking, it’s being performed outside of the context of the cell. What we’re doing is taking the cells, grinding them into pieces, isolating the protein constituents and then asking how they work back in their native habitat.
That’s like going to Taco Bell to learn about Mexican food. We’re missing the big picture. What we really wish to do is visualize viral integration unfold in real time inside of a live cell with atomic resolution. This will enable us to define exactly how, when and where integration occurs, facilitating the discovery of novel ways to interrupt the process.
The Holy Grail is really about visualizing and describing the molecular world in the context of a crowded native cellular environment. This is extremely challenging. There’re many problems that we’ll need to overcome.
Most cells are too dense for electrons to penetrate. Our field has been developing tools to thin them down and hone in on the event of interest. There’s a myriad of technical problems like accuracy, reproducibility, contamination, needing better detectors and unconventional imaging strategies. We will need new algorithms, faster computers, smarter automation and even a bit of AI to annotate everything that we see.
If we can solve the technical challenges, then we’ll reveal the very first glimpse of the viral molecular protein arsenal that continues to devastate the world population. We’ll be able to visualize the battle scenes in HD. By seeing biology at a molecular level, we can define new therapeutic blueprints that will allow us to put in further molecular protections to change the course of HIV therapy.
It’s a complex problem. There is no cure. So, let’s not stop there. Beyond viral integration, we will open up a whole new world of exploration in our own cells, about which we know surprisingly little at the molecular level.
The vast majority of the molecular organization of the cell is uncharted territory. Visualizing the molecular processes associated with infection is just one of the many scientific areas that we need to better understand to chart a path to a cure.
Sun Tzu finally said, “Know yourself and know the enemy. A thousand battles, a thousand victories.” We don’t need to yield to viral onslaught. Let’s chart out the molecular landscape of infection so we can develop our own protective arsenal to stop this and thwart the next deadly outbreak.