Identification of a Second Major Site for CD46 Binding in the Hemagglutinin Protein from a Laboratory Strain of Measles Virus (MV): Potential Consequences for Wild-Type MV Infection

Abstract

Natural or wild-type (wt) measles virus (MV) infection in vivo which is restricted to humans and certain monkeys represents an enigma in terms of receptor usage. Although wt MV is known to use the protein SLAM (CD150) as a cell receptor, many human tissues, including respiratory epithelium in which the infection initiates, are SLAM negative. These tissues are CD46 positive, but wt MV strains, unlike vaccinal and laboratory MV strains, are not thought to use CD46 as a receptor. We have identified a novel CD46 binding site at residues S548 and F549, in the hemagglutinin (H) protein from a laboratory MV strain, which is also present in wt H proteins. Our results suggest that although wt MV interacts with SLAM with high affinity, it also possesses the capacity to interact with CD46 with low affinity.Measles virus (MV), a member of the Morbillivirus genus in the Paramyxoviridae family in the Mononegavirales order, is responsible for at least 1 million infant deaths each year worldwide. This high mortality is not due directly to MV infection but to a transient, profound immunosuppression induced by the virus which allows the propagation of pathogenic secondary infections. The present MV vaccines are derived from the Edmonston strain, which was isolated in 1954 using primary human kidney cells and then attenuated by further passaging on human amnion cells and chicken embryo fibroblasts (9). Edmonston was also attenuated by adaptation to African green monkey kidney (also known as Vero) cells, a procedure that became standard practice for the isolation of MV laboratory strains which phenotypically resemble vaccine strains. In 1990, however, Kobune et al. (14) showed that MV strains could be rapidly isolated using the Epstein-Barr virus-transformed simian B-lymphoblastic cell line B95-8 and its adherent subline B95a. Such strains are not attenuated but pathogenic and resemble circulating wild-type (wt) MV strains.MV possesses two glycoproteins in its envelope: the hemagglutinin (H) protein (MVH) and the fusion protein (MVF). MVH is responsible for attachment to the cellular receptor, whereas MVF mediates the fusion of the viral and host cell membranes (34). MV fusion has been shown to depend upon the coexpression of the two glycoproteins (36) It is believed that the fusion helper function of the H protein depends upon a specific physical interaction with the MVF which is mediated by the cysteine-rich domain of the latter protein (35). Nearly 10 years ago it was shown that two laboratory strains of MV use the protein CD46, a member of the regulators of complement activation superfamily, as a cellular receptor (5, 23). Although the MVH-CD46 interaction is probably conformational and many amino acids on MVH contribute, it was found that a tyrosine residue at position 481 of MVH plays a crucial role: mutation of this residue leads to abrogation of the interaction with CD46 (1,11,17). The observation that residue 481 is asparagine in most if not all MVH proteins from wt MV strains and the finding that anti-CD46 antibodies did...

Multiple Amino Acid Substitutions in Hemagglutinin Are Necessary for Wild-Type MeaslesVirus To Acquire the Ability To Use Receptor CD46 Efficiently

“…Residues at positions 390 and 416 are situated in ␤-sheet 3, those at positions 446 and 481 are in ␤-sheet 4, and those at positions 484 and 492 are in the connecting loop between ␤-sheets 4 and 5. Thus, CD46-relevant residues identified by previous and present studies are localized over a wide range of domains in the H protein (3,24,33,55). It is possible that some of the residues identified are involved in direct CD46 binding, whereas others affect the structure of the H protein such that its binding site can interact with CD46 efficiently.…”

“…Interestingly, the reference strain of genotype D2 (Johannesburg.SOA/88/1) has an asparagine at position 484 of the H protein in addition to a tyrosine at position 481, whereas that of genotype E (Goettingen.DEU/71 "Braxator") possesses glycine at positions 492 and 546 of the H protein (57). It is known that S546G substitution in the H protein, like the N481Y substitution, allows MV to use CD46; indeed, some Vero cellgrown MVs have an S546G substitution in the H protein instead of N481Y (13,14,21,24,27,32,36,37,43,46,56). Thus, these two reference strains seem to have the ability to use CD46 efficiently.…”

“…A series of H protein mutants were examined for SLAM-or CD46-dependent fusion-inducing activity, and the residues important for the interaction with SLAM and CD46 were identified (23,24,55). Those studies showed that CD46-relevant residues are mainly located in ␤-sheet 4 (positions 428, 431, 451, 452, 464, and 481) and the connecting loop between ␤-sheets 4 and 5 (positions 486 and 487) of the ectodomain, and some residues might reside in ␤-sheet 5 (positions 527, 546, 548, and 549).…”

Selectively Receptor-Blind Measles Viruses: Identification of Residues Necessary for SLAM- or CD46-Induced Fusion and Their Localization on a New Hemagglutinin Structural Model

“…Nevertheless, a recombinant virus with a wild-type H-protein N481 residue does propagate in Vero cells (31), suggesting either that Y481 is not an absolute requirement for CD46 binding or that a second receptor exists on Vero cells (25). Other studies identified amino acids 473 to 477 as another site required for CD46 interactions (53) and amino acids 548 to 549 as a site required for CD46 downregulation (38). Moreover, residue 546 is at variance between different strains, and it has been suggested that it may play a role in virus adaptation to different growth conditions (30,62,75).…”

Identification of Hendra Virus G Glycoprotein Residues That Are Critical for Receptor Binding

“…The domains of MeV H that engage CD46 and CD150 have been modeled to lie along the top of the globular head, forming conformation-dependent and overlapping binding sites composed primarily of residues in beta sheets 4 and 5 (22,24,25,36). The structure of the henipavirus G has yet to be determined; however, the amino acid sequences of both HeV and NiV G proteins can accommodate a six-bladed beta-propeller structural model similar to other paramyxovirus H or HN glycoproteins (38,40).…”