Viruses evolve due to constantly changing survival environments. Most importantly, the natural evolution of the viruses tends to be faster than their hosts, such as humans. That makes us vulnerable to new virus variants as we are facing currently with the recent outbreak of a new coronavirus, SARS-CoV-2, that sadly led to worldwide large number of lost human lives. The spread of the virus resulted in unprecedented health and socio-economic crisis. Enteroviruses are some of the most common human viral pathogens, and, similarly to coronaviruses, enteroviruses are positive-sense, single-stranded RNA viruses capable of causing a wide spectrum of illness, including hand-foot-and-mouth disease, conjunctivitis, hepatitis, myocarditis, meningitis, encephalitis. In 2014, outbreaks of severe respiratory illness associated with enterovirus D68 occurred among children in the North America. Except poliovirus, we lack vaccines and antivirals therapies for many of the enteroviruses. This is partly due to high mutation rates of enteroviruses and likelihood of acquired resistance. That is why there is an urge to understand the mechanisms these viruses are using when infecting human cells, how they cause disease, or how our bodies react to infections. To answer such an important biological question, one must provide a strong biological answer and bridge the space between fundamental research and applicable research in designing new therapies. In many cases, structures of isolated enterovirus particles been studied in detail. However, the structure and mechanism of their replication complexes, the main intracellular manifestation of the viruses, is still poorly understood. The approach I propose here will be a unique attempt to reveal the structural basis of how cellular membranes are disrupted and reshaped to support enterovirus infection. Using novel cryo-focused ion beam milling technology combined with cryo-electron tomography, we could visualize and identify molecular assemblies in enterovirus-infected cells, not visible in conventional EM. This ultimately helped us to characterize further how the virus hijacks the host cellular machinery for its optimum replication. The main impact of this project will thus be structural and molecular insights on how enteroviral proteins can coordinate a remodeling of cellular membranes and create a localized viral genome factory. This will be among the first high-resolution structural study of viral replication complexes in cells and will thus serve as a paradigm for molecular understanding of pathogen-induced membrane remodeling. Thus, the proposed project has the potential to provide solid scientific benefits and potentially also new concepts for therapeutics.
