Disruption of the Dimer-Dimer Interaction of the Mumps Virus Attachment Protein Head Domain, Aided by an Anion Located at the Interface, Compromises Membrane Fusion Triggering

Abstract

Mumps virus (MuV), an enveloped negative-strand RNA virus belonging to the family Paramyxoviridae, enters the host cell through membrane fusion mediated by two viral envelope proteins, an attachment protein hemagglutinin-neuraminidase (MuV-HN) and a fusion (F) protein. However, how the binding of MuV-HN to glycan receptors triggers membrane fusion is not well understood. The crystal structure of the MuV-HN head domain forms a tetramer (dimer of dimers) like other paramyxovirus attachment proteins. In the structure, a sulfate ion (SO42−) was found at the interface between two dimers, which may be replaced by a hydrogen phosphate ion (HPO42−) under physiological conditions. The anion is captured by the side chain of a positively charged arginine residue at position 139 of one monomer each from both dimers. Substitution of alanine or lysine for arginine at this position compromised the fusion support activity of MuV-HN without affecting its cell surface expression, glycan-receptor binding, and interaction with the F protein. Furthermore, the substitution appeared to affect the tetramer formation of the head domain as revealed by blue native-PAGE analysis. These results, together with our previous similar findings with the measles virus attachment protein head domain, suggest that the dimer-dimer interaction within the tetramer may play an important role in triggering membrane fusion during paramyxovirus entry. IMPORTANCE Despite the use of effective live vaccines, mumps outbreaks still occur worldwide. Mumps virus (MuV) infection typically causes flu-like symptoms and parotid gland swelling but sometimes leads to orchitis, oophoritis, and neurological complications, such as meningitis, encephalitis, and deafness. MuV enters the host cell through membrane fusion mediated by two viral proteins, a receptor-binding attachment protein, and a fusion protein, but its detailed mechanism is not fully understood. In this study, we show that the tetramer (dimer of dimers) formation of the MuV attachment protein head domain is supported by an anion located at the interface between two dimers and that the dimer-dimer interaction plays an important role in triggering the activation of the fusion protein and causing membrane fusion. These results not only further our understanding of MuV entry but provide useful information about a possible target for antiviral drugs.

Evaluation of the Interactions between Mumps Virus and Guinea Pig

“…Since it was shown that a trisaccharide containing α2,3-linked sialic acid at the cell surface acts as a receptor for MuV ( 2 , 3 ), the expression of SAs at the surface of guinea pig primary cell cultures from brain, testicles, lung, and kidney was measured. SAs were released by treatment with α2,3-sialidase (removes α2,3-sialyl groups) and Arthrobacter ureafaciens sialidase (removes the following sialyl groups at rate α2,6→α2,3→α2,8-linkage).…”

“…Mumps orthorubulavirus (MuV) belongs to the genus Orthorubulavirus of the family Paramyxoviridae , which includes enveloped, nonsegmented, negative-strand RNA viruses ( 1 ). Two MuV envelope glycoproteins, hemagglutinin-neuraminidase (HN) and fusion (F) protein, are involved in receptor binding and mediate membrane fusion with target cells ( 2 , 3 ). HN specifically recognizes sialic acid (SA)-containing glycoconjugate structures present on the host cells, preferring unbranched α2,3-sialylated glycans, which therefore play a key role in virus entry into host cells and infectivity ( 4 ).…”

Exploring the Mumps Virus Glycoproteins: A Review

“…The helix-bundled stalk of the HN protein is thought to play a critical role in the activation of the F protein. The dimer-dimer interactions found in the tetrameric head domain of the HN protein seem to be important in the role of the HN protein triggering the F protein for fusion [37,38].…”

Mumps Orchitis: Clinical Aspects and Mechanisms

“…A previous study demonstrated that MuV used a trisaccharide containing α2,3-linked sialic acid on the cell surface as a receptor that interacts with MuV attachment protein HN ( 61 ). The MuV-HN-receptor interaction triggers the activation of the F protein, causing fusion of the viral envelope with the plasma membrane and allowing cell entry ( 62 ). A very recent study has confirmed the presence of sialic acid on the surface of Sertoli cells (SCs) and Leydig cells (LCs) ( 63 ).…”