In order to examine functions of the hemagglutinin-neuraminidase (HN) protein that quantitatively influence fusion promotion, human parainfluenza virus 3 (HPIV3) variants with alterations in HN were studied. The variant HNs have mutations that affect either receptor binding avidity, neuraminidase activity, or fusion protein (F) activation. Neuraminidase activity was regulated by manipulation of temperature and pH. To dissect the specific contribution of neuraminidase to triggering, two variant HNs that are triggering-defective due to a mutation in the HN stalk were evaluated. One of these variants has, in addition, a mutation in the globular head that renders it neuraminidase dead, while the HN with the stalk mutation alone has 30% of wt neuraminidase. While the variant without neuraminidase activity triggered F effectively at 37°C irrespective of pH, the variant possessing effective neuraminidase activity completely failed to activate F at pH 5.7 and was capable of only minimal triggering activity even at pH 8.0. These results demonstrate that neuraminidase activity impacts the extent of HPIV3-mediated fusion by releasing HN from contact with receptor. Any particular HNs competence to promote F-mediated fusion depends on the balance between its inherent F-triggering efficacy and its receptor-attachment regulatory functions (binding and receptor cleavage).Human parainfluenza virus 3 (HPIV3), like other paramyxoviruses, possesses two envelope proteins directly involved in viral entry and cytopathology. Hemagglutinin-neuraminidase (HN), by binding to sialic acid-containing cellular surface receptors, brings the viral envelope in proximity to the plasma membrane. Fusion protein (F) is the active mediator of fusion between the viral and cell membranes, which leads to release of the viral nucleocapsid into the cytoplasm. In order to mediate fusion, F must be present in its cleaved state and then undergo a second activation step to assume its fusion-ready conformation. It has been shown that HN plays an essential role in the fusion process, and one proposed role of HN is to drive the final conformational change in F that renders it fusion active (30). Fusion of the virus with the target cell, which permits synthesis of viral macromolecules and expression of viral envelope proteins on the host cell's surface, is followed by two processes that lead to the spread of infection and HPIV3 cytopathogenesis. One is the assembly and budding of new virions that then infect distant as well as neighboring cells. Second, by virtue of HN and F expressed on its surface, an infected cell can fuse with adjacent uninfected cells, forming syncytia. In the spread of infection via budding, HNЈs receptordestroying neuraminidase activity plays the essential role of preventing progeny virions from remaining aggregated on the infected cell's surface rather than spreading to additional cells (29). For syncytium formation, no direct role has yet been found for HNЈs neuraminidase activity, although the magnitude of the receptor-cleaving activity...

Hendra virus (HeV) is a recently identified paramyxovirus that is fatal in humans and could be used as an agent of bioterrorism. The HeV receptor-binding protein (G) is required in order for the fusion protein (F) to mediate fusion, and analysis of the triggering/activation of HeV F by G should lead to strategies for interfering with this key step in viral entry. HeV F, once triggered by the receptor-bound G, by analogy with other paramyxovirus F proteins, undergoes multistep conformational changes leading to a six-helix bundle (6HB) structure that accomplishes fusion of the viral and cellular membranes. The ectodomain of paramyxovirus F proteins contains two conserved heptad repeat regions (HRN and HRC) near the fusion peptide and the transmembrane domains, respectively. Peptides derived from the HRN and HRC regions of F are proposed to inhibit fusion by preventing F, after the initial triggering step, from forming the 6HB structure that is required for fusion. HeV peptides have previously been found to be effective at inhibiting HeV fusion. However, we found that a human parainfluenza virus 3 F-peptide is more effective at inhibiting HeV fusion than the comparable HeV-derived peptide.