Functional relevance of the transmembrane domain and cytoplasmic tail of the pseudorabies virus glycoprotein H for membrane fusion

Vallbracht, Melina GND; Fuchs, Walter GND; Klupp, Barbara GND; Mettenleiter, Thomas C. GND

Herpesvirus membrane fusion depends on the core fusion machinery comprising glycoproteins (g)B and gH/gL. Although gB structurally resembles autonomous class III fusion proteins, it strictly depends on gH/gL to drive membrane fusion. Whether the gH/gL complex needs to be membrane-anchored to fulfill its function and which role the gH cytoplasmic (CD) and transmembrane domains (TMD) play in fusion is unclear. While the gH CD and TMD play an important role during infection, soluble gH/gL of herpes simplex virus 1 (HSV-1) seems to be sufficient to mediate cell-cell fusion in transient assays arguing against an essential contribution of the CD and TMD. To shed more light on this apparent discrepancy we investigated the role of the CD and TMD of the related alphaherpesvirus pseudorabies virus (PrV) gH. For this purpose, we expressed C-terminally truncated and soluble gH, and substituted the TMD by a glycosylphosphatidylinositol (gpi)-anchor. We also generated chimeras containing the TMD and/or CD of PrV gD or HSV-1 gH. Proteins were characterized in cell-based fusion assays and during virus infection. Although truncation of the CD resulted in decreased membrane fusion activity, the mutant proteins still supported replication of gH-negative PrV, indicating that the PrV gH CD is dispensable for viral replication. In contrast, PrV gH lacking the TMD, membrane-anchored via a lipid-linker, or comprising the PrV gD TMD were non-functional highlighting the essential role of the gH TMD for function. Interestingly, despite low sequence identity, the HSV-1 gH TMD could substitute for the PrV gH TMD pointing to functional conservation.

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Vallbracht, Melina / Fuchs, Walter / Klupp, Barbara / et al: Functional relevance of the transmembrane domain and cytoplasmic tail of the pseudorabies virus glycoprotein H for membrane fusion. 2018.

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