Dynamic Interaction of the Measles Virus Hemagglutinin with Its Receptor Signaling Lymphocytic Activation Molecule (SLAM, CD150)

Abstract

The interaction of measles virus with its receptor signaling lymphocytic activation molecule (SLAM) controls cell entry and governs tropism. We predicted potential interface areas of the measles virus attachment protein hemagglutinin to begin the investigation. We then assessed the relevance of individual amino acids located in these areas for SLAM-binding and SLAM-dependent membrane fusion, as measured by surface plasmon resonance and receptor-specific fusion assays, respectively. These studies identified one hemagglutinin protein residue, isoleucine 194, which is essential for primary binding. The crystal structure of the hemagglutinin-protein localizes Ile-194 at the interface of propeller blades 5 and 6, and our data indicate that a small aliphatic side chain of residue 194 stabilizes a protein conformation conducive to binding. In contrast, a quartet of residues previously shown to sustain SLAM-dependent fusion is not involved in binding. Instead, our data prove that after binding, this quartet of residues on propeller blade 5 conducts conformational changes that are receptor-specific. Our study sets a structure-based stage for understanding how the SLAM-elicited conformational changes travel through the H-protein ectodomain before triggering fusion protein unfolding and membrane fusion.Measles is a leading cause of childhood morbidity and mortality in developing countries (1). The immune suppression that accompanies infection with wild-type measles viruses (MV) 2 exposes individuals to secondary infections causing most of the fatalities associated with the disease (2). Live attenuated MV vaccines have effectively reduced the morbidity and mortality of measles world-wide (3). The difference in pathology between the wild-type and vaccine strains is in part explained by the receptors used for cell entry. Wild-type MV uses the signaling lymphocyte activation molecule (SLAM, CD150) (4 -7), while the vaccine strain has gained the ability to also use the ubiquitous protein CD46 (8 -10). SLAM-targeting accounts for lymphotropism and is central to understanding MV tropism and disease progression.Receptor attachment of MV and the other morbilliviruses is mediated by hemagglutinin (H) homodimers. Unlike other members of the Paramyxoviridae, the morbillivirus H-proteins do not have neuraminidase activity. Upon receptor attachment, H induces conformational changes in the trimeric fusion (F) protein, resulting in the fusion of viral and cellular membranes (11,12). MV H is a 617-amino acid type II transmembrane glycoprotein, which is comprised of an N-terminal cytoplasmic tail, a membrane-spanning domain, and an extracellular stalk region connected to a C-terminal globular head (13).Previously, we generated a three-dimensional model of the extracellular domain of the MV H-protein based on the crystal structure of Newcastle Disease Virus (NDV) HN (hemagglutinin neuraminidase) protein (sequence identity of 12%), a related paramyxovirus (14,15). This model predicts that the MV H ectodomain has a 6-blade propeller structure....

Pseudotyping lentiviral vectors with the wild-type measles virus glycoproteins improves titer and selectivity

“…To further show that cell entry of the MV WT -HIV particles was SLAM-independent, we generated vector particles with the 194A mutant of the H WT protein, which interferes with SLAM binding. 22 These particles transduced small airway epithelial cells with high efficiency (11% GFP + cells) but were unable to transduce Raji cells (data not shown).…”

“…As the H residues contacting SLAM are different from those contacting EpR, 7,22 EpR-specific vectors that are SLAMblind can be generated. As the EpR is expressed basolaterally, gene transfer to lung and other epithelia upon systemic application may become feasible with this vector.…”

Probing the Spatial Organization of Measles Virus Fusion Complexes

“…Figure 2C exemplifies that, following binding of this sSLAM-IgG construct to H-expressing cells by flow cytometry, the ratios of SLAM-specific mean fluorescence intensity to H-specific mean fluorescence intensity provide an assessment of receptor (SLAM) binding competence. For a control, an I194S mutant (H-I194S) that reportedly is unable to bind to SLAM was assessed (48). This construct returned a SLAM-to-H signal ratio of 0.03, compared to 1.07 obtained for standard H (Fig.…”

“…If H stalks enjoy some rotational flexibility and residues in stalk section 84 to 118 (presumably residues 111, 114, and 118) engage in short-range contacts for F triggering, how is receptor binding by the H head domain (48,71) signaled to the contact zone despite the presence of a long (41-residue) stalk insertion? Interestingly, no major conformational changes between crystal structures of PIV5 HN, HPIV3 HN, and henipavirus G, solved alone or in complex with their receptors, were observed (8,9,74).…”

Measles virus blind to its epithelial cell receptor remains virulent in rhesus monkeys but cannot cross the airway epithelium and is not shed

“…The 3 EpR-relevant residues (red) define a small triangle located slightly above the equator. I194, the residue governing SLAM-binding (27), and the 4 residues involved in SLAM-dependent conformational changes (purple) (16) are located centrally on the top of each dimer. Most CD46-relevant residues (yellow) are in the bottom half.…”