HIV-1, like other viruses, lacks the machinery to produce its own proteins and must rely on the host cell to translate its genetic instructions. After entering host cells, it seizes control of the translation process, which converts messenger ribonucleic acid (mRNA) into proteins. "In this study, we combined ribosome profiling, RNA sequencing and RNA structural probing to map the viral and host translational landscape and pausing during replication of the virus in unprecedented detail," says corresponding author Neva Caliskan. She is a former group leader at the Helmholtz Institute for RNA-based Infection Research (HIRI) in Würzburg, a site of the Braunschweig Helmholtz Centre for Infection Research (HZI) in cooperation with the Julius-Maximilians-Universität Würzburg (JMU), and is currently the Director of the Department of Biochemistry III at the University of Regensburg.
Cheat Codes of Viral Translation
One of the key findings was the discovery of previously unrecognized elements in HIV-1 RNA called upstream open reading frames (uORFs) and internal open reading frames (iORFs). These "hidden gene fragments" may play a crucial role in fine-tuning the production of viral proteins as well as the interaction with the host immune system. "For instance, uORFs and iORFs can act as regulators, ensuring precise timing and levels of protein synthesis", explains Anuja Kibe, a postdoctoral researcher at the HIRI and first author of the study, which was published in the journal Nature Structural and Molecular Biology.
Another important discovery was an intricate RNA structure near the critical "frameshift site" in the viral genome. This frameshift site is essential for the virus to produce the correct proportions of two key proteins, Gag and Gag-Pol, which are necessary for assembling infectious particles and replication of HIV-1. The researchers demonstrated that this extended RNA fold not only promotes ribosome collisions upstream of the site—a mechanism that appears to regulate translation—but also maintains the frameshifting efficiency. "Our team also showed that targeting this RNA structure with antisense molecules could significantly reduce frameshift efficiency by nearly 40 percent, offering a promising new avenue for antiviral drug development", reports Caliskan.
A Game of Priorities
Redmond Smyth, a former Helmholtz Young Investigator Group Leader at the HIRI and currently a group leader at the Centre National de Recherche Scientifique (CNRS) in Strasbourg, France, mentions, "Interestingly, our analysis revealed that, while HIV-1 mRNAs are translated efficiently throughout infection, the virus suppresses the protein production of the host, particularly at the translation initiation stage." This allows HIV-1 to prioritize its own needs while effectively stalling the host defense mechanisms. Thus, the virus can manipulate the host cell machinery in ways that remain robust even under stress conditions.
More Than Traffic Jams
The researchers also observed that ribosomes collide at specific regions of the viral RNA, particularly upstream of the frameshift site. "These collisions are not accidental but are instead tightly regulated pauses that may influence how ribosomes interact with downstream RNA structures," says Florian Erhard, study co-author and Chair of Computational Immunology at the University of Regensburg.
Overall, the study provides not only a detailed map of the translational landscape of HIV-1 infected cells but also a wealth of potential targets for therapeutic intervention. The identification of RNA structures and genetic elements critical for viral replication highlights new opportunities for the development of drugs aimed at disrupting these processes. "By understanding how the virus cleverly manipulates our cells, these discoveries will bring us closer to innovative treatments that could one day turn tables and outsmart the virus itself," Caliskan adds.
Funding
The study was primarily funded by the European Research Council Starting Grant to Neva Caliskan (Grant No. 948636) and by the Helmholtz Association. Redmond Smyth received funding from the Helmholtz Young Investigator Grant VH-NG-1347 and the Center for Structural Biology of HIV-1 RNA U54 AI170660.
Helmholtz Institute for RNA-based Infection Research
The Helmholtz Institute for RNA-based Infection Research (HIRI) is the first institution of its kind worldwide to combine ribonucleic acid (RNA) research with infection biology. Based on novel findings from its strong basic research program, the institute's long-term goal is to develop innovative therapeutic approaches to better diagnose and treat human infections. HIRI is a site of the Braunschweig Helmholtz Centre for Infection Research (HZI) in cooperation with the Julius-Maximilians-Universität Würzburg (JMU) and is located on the Würzburg Medical Campus. More information at www.helmholtz-hiri.de .
Helmholtz Centre for Infection Research
Scientists at the Helmholtz Centre for Infection Research (HZI) in Braunschweig and its other sites in Germany are engaged in the study of bacterial and viral infections and the body's defence mechanisms. They have a profound expertise in natural compound research and its exploitation as a valuable source for novel anti-infectives. As member of the Helmholtz Association and the German Center for Infection Research (DZIF) the HZI performs translational research laying the ground for the development of new treatments and vaccines against infectious diseases. https://www.helmholtz-hzi.de/en
