The Role of the Nipah Virus G Head and Stalk Regions in Modulating Viral Fusion

Thesis (Ph.D.), College of Veterinary Medicine, Washington State UniversityNipah virus (NiV) is an emerging zoonotic virus within the paramyxovirus family that causes severe neurological and respiratory disease in its victims and has mortality rates in humans of up to one hundred percent. Membrane fusion is critically important for NiV spread in the form of both viral-cell fusion for viral entry and cell-cell fusion (syncytia formation) for cell to cell spread. The fusion process is facilitated by two glycoproteins on the viral membrane: the attachment protein G and the fusion protein F. Binding of G to the cellular receptor ephrinB2 or ephrinB3 triggers F to undergo a conformational cascade that results in membrane fusion. Though a general mechanism of fusion is somewhat understood, the specifics of this process are still largely unknown. Here we aimed to elucidate some critical steps in the fusion process, with the hope that better understanding this fusion process will pave the way for future preventative strategies against NiV spread. NiV G consists of an N terminal cytoplasmic tail, followed by a transmembrane region, followed by an extracellular stalk and globular head domain on the C terminal. In chapter one we showed that two conformational changes occur in the NiV G head upon receptor binding that are critical for fusion triggering. In chapter two we investigated the effects of O-glycans, a severely understudied protein modification process, in the NiV G stalk, determining that O-glycans play roles in cell-cell fusion, F and G interactions, G conformations, viral entry, and F incorporation into virions. In chapter three we developed a novel flow cytometry method for studying interactions of membrane proteins and used this strategy to show a bi-dentate interaction whereby both the stalk and head domains of NiV G are interacting with NiV F, a novel finding for the paramyxoviruses. Taken all together these findings have helped further elucidate steps and regions within NiV G that are important in the fusion triggering process.College of Veterinary Medicine, Washington State Universit