Studying Factors that Contribute to Uncoating of Enteroviruses

Abstract

Enteroviruses are small, non-enveloped viruses with a positive sense single stranded RNA genome. They cause different diseases in humans, usually with symptoms of common cold, but also more severe acute and chronic infections such as encephalitis and type 1 diabetes. Although the structure and infection pathway of many entero-viruses is rather well-known, many important details remain unresolved. For some enteroviruses receptor binding or low pH has been shown to convert them from an intact to an intermediate particle, which is needed for successful infection. However, B-species enteroviruses do not rely on the same factors e.g. low pH for efficient infec-tion. Furthermore, the decisive factor releasing the enterovirus genome, is still un-known. Learning these missing factors is important for understanding the virus in-fection in more detail, that in turn provides basis for developing antiviral strategies. This study concentrates on two B-species enteroviruses, echovirus 1 and cox-sackievirus A9, aiming to resolve if physiological factors, serum albumin and ion changes during the virus infection, trigger the virus transformation from intact to altered particles, and possibly further to genome release. We found that both factors contribute to formation of an intermediate particle of both viruses. Furthermore, spe-cific changes in the ionic milieu led to the final genome release. The studied factors resulted in rather similar changes in their cryo-EM structures that are found for other enteroviruses primed using factors such as heat, low pH and receptor binding. However, we found that priming the coxsackievirus A9 using ion changes or albu-min resulted in slightly different changes in virus capsid proteins: albumin resulted in more stable virus intermediate particle, whereas ions altered the virus capsid into a stage closer to the genome release. In the third part of this study, the aim was to develop a novel tool for live cell imaging of enterovirus entry and uncoating. We first solved the structure of intercalating RNA dye SYBR green II, modified it for better fluorescent properties and different binding capabilities, and finally verified its supe-riority in virus uncoating assays.