DNA transport

The genome of HIV-1 embarks on a multistep journey when a cell is infected. In the infectious virion, the genome is carried in two strands of RNA, protected inside a cone-shaped capsid core. This core is injected into the cell. Enzymes in the core then make use of the ready supply of nucleotides in the infected cell, building a DNA copy of the genome. Finally, the capsid delivers this DNA genome to the nucleus, where it is integrated into the cell’s own genome. As part of this process, the capsid is thought to pass through the nuclear pore complex, the gatekeeper between the cell’s cytoplasm and nucleus. Whether the capsid passes through the nuclear pore complex intact or is broken apart during entry remains a long standing question in HIV-1 biology.

Coarse-grain simulation

B-HIVE researchers have used coarse-grained simulation to explore the molecular details of the passage of HIV-1 capsid through the nuclear pore complex. In coarse-grained simulation, simplified models are designed that capture the physical properties of each of the molecules. In this way, very complex systems may be studied while still ensuring that the motions and interactions of the individual proteins are consistent with more detailed atomic studies. This type of coarse-graining is essential for study of capsid and the nuclear pore complex, since each is composed of over a thousand proteins. Studying this process at atomic detail is beyond the capabilities of even the most sophisticated supercomputers currently available.

What this tells us about HIV

The details of how HIV-1 capsid gets through the pore and releases the viral DNA are still largely a mystery. This study allows us to explore the process in detail, identifying weak points in the virus that would provide avenues for therapeutic intervention. For example, the simulations identify stripes of capsid proteins that are particularly stressed when the capsid is passing through the pore. Current drug design efforts are looking for ways to strengthen the connections between these capsid proteins, with the intention of blocking release of the viral genome into the nucleus.

Simulations explore two ways that HIV-1 capsid approaches the nuclear pore complex. If the pointed end enters first (top), it is able to pass through, but the opposite orientation (bottom) is blocked. The study also explored capsids with different shapes and different states of the nuclear pore complex.

Simulation revealed that stripes of capsid protein (red) undergo increased stress as the capsid core passes through the nuclear pore complex.

Arpa Hudait

How did you get interested in science?

I grew up in an industrial and manufacturing hub in India, surrounded by engineering and science professionals working in the mining, chemical, and power industries. In addition to education at school, I was lucky to regularly visit these industrial, manufacturing, and scientific facilities as part of school tours and outreach events (a rare privilege in a lower income and industrially developing nation). I had a keen interest in both chemical processes and fundamental principles of physics. I am a visual learner, and can vividly visualize in mind anything I read. Hence, the idea of simulating the motions of a small molecule or protein in a computer and watching the movies was fascinating to me. That led me to pursue research in computational chemistry during my undergraduate. I moved to the beautiful Salt Lake City to pursue my graduate studies at the University of Utah. I continued working on computational chemistry and biophysics, where I modeled ice formation in atmospheric aerosols and also studied ice-binding proteins found in organisms surviving cold conditions. For my postdoctoral research, I moved to the University of Chicago to work in the group of Prof. Greg Voth. At the University of Chicago, I focused on modeling processes in the HIV-1 life cycle.

Tell us about the lab where you did this work.

I did this work in the group of Prof. Greg Voth at the University of Chicago. Computer modeling of biomolecular complexes is essential to provide a mechanistic picture of cellular processes. Typically, simulating these processes in atomistic resolution is infeasible due to high computational costs. Therefore, coarse-grained models, which are simplified versions of atomically detailed models, are essential. The Voth group in the past 20 years has been the leader in surmounting the critical challenges of developing effective techniques to systematically derive coarse-grained models of high fidelity.

I had the opportunity to join the Voth group for my postdoctoral research. It was the perfect opportunity to train in the computational methodologies of coarse-graining and expand my expertise. This move to the Voth group at the University of Chicago was perfect to apply these methodologies to develop coarse-grained models of complex biomolecular systems, specifically of HIV-1 proteins. Initially, at the Voth group, I investigated ESCRT protein assembly. After joining B-HIVE, my research had been focused on modeling nuclear entry of HIV-1 capsid, and capsid-host interactions. 

What were the biggest challenges with this study?

Nuclear pore complex (NPC) is one of the largest macromolecular complexes in the cell, consisting of over 1000 proteins. Therefore, modeling this complex is extremely challenging due to its size, multiple unique protein-protein heteromultimeric assembly interfaces, and protein-membrane interaction sites. Additionally, the capsid complex containing the viral RNA consists of 1000 proteins and interacts with the NPC mediated by interactions with the heterogeneous sieve-like environment of disordered chains rich in phenylalanine and glycine residues. Therefore, we first developed a coarse-grained model of the human NPC from the recently released high-resolution structural model. We employed a “divide and conquer” strategy and simulated smaller protein complexes (instead of the full NPC) separately in detailed atomistic resolution. From these atomistic simulation configurations, we systematically derived the coarse-grained model and interactions of the NPC. We then assembled a composite membrane-embedded model of the NPC and simulated capsid docking.

To understand the factors that regulate the successful passage of HIV-1 capsid at the NPC, we simulated different capsid shapes and orientation of approach. Cone-shaped capsid is the most prevalent form of the capsid. Additionally, tubular and ellipsoid shaped capsids are also observed in imaging studies in infected cells. We simulated the docking of the cone-shaped capsid when the narrow tip and wide tip of the capsid approached the pore. We find that when the capsid approaches from the narrow end, it successfully establishes contact with the capsid-binding phenylalanine glycine chains. Due to the increasing diameter of the capsid, the pore dilates to accommodate the capsid. These findings demonstrate that the nuclear pore is a pliable complex capable of regulating nucleocytoplasmic transport. We discovered that capsids docked at the nuclear pore develop correlated striated patterns in the lattice. These patterns are a measure of capsid flexibility and are formed as a consequence of the stress from the confinement at the pore. Expansion of the internal genome mimicking early stages of reverse transcription further accentuates these weak points. Despite the structural weakness, we demonstrate that the capsid remains intact during docking at the nuclear pore. To summarize, this is a challenging study to perform from the perspective of computational modeling due to the incredible complexity of the system. Our study provides an example of how computer simulations using quantitatively developed models can be used to improve the mechanistic understanding of the dynamics of complex cellular processes.

What are you working on now?

​​I am currently continuing my research on capsid entry into the nuclear pore complex, particularly focusing on therapeutic intervention. Previously, my work has demonstrated that there are weak points in the capsid lattice. Essentially, the capsid weak points are a measure of the capsid molecular flexibility. I am currently simulating how the antiviral drug Lenacapavir targets these weak points to pinpoint the mode of action of drugs with the capsid protein. The key goal of my current research in B-HIVE is to understand whether modulating the intrinsic capsid molecular flexibility through antiviral binding can rupture the capsid during nuclear entry and limit infection.