Grants and Contracts Details
The paramyxovirus human metapneumovirus (HMPV) is still a major cause of viral respiratory disease in infants, the elderly, and in immunocompromised patients, second only to respiratory syncytial virus (RSV). Several studies have shown that preexisting cardiopulmonary disease is one of the major risk factors leading to hospitalizations from severe HMPV infection in children and adults. HMPV has a fusion (F) protein on its surface that, upon triggering, fuses the viral and cellular membranes during viral entry. Most paramyxovirus F proteins can be triggered at neutral pH. However, we have shown that low pH is a strong stimulus for HMPV F protein triggering. Furthermore, we have shown that mutations in the HRB-linker domain of the F protein can abolish or greatly diminish the stimulation provided by low pH. These observations prompted us to further investigate the entry mechanism of this virus. Initial data on HMPV infection showed that viral entry can be partially blocked by inhibitors of endosomal acidification, an observation consistent with the low pH stimulation observed in our F protein studies. Chlorpromazine, an inhibitor of clathrin-mediated endocytosis, was also able to reduce HMPV infection. Furthermore, dynasore, a small-molecule inhibitor of dynamin, was able to block HMPV infection by as much as 90%. These results are very interesting as it is generally believed that paramyxoviruses enter the cell at the plasma membrane, where the pH is neutral. Our data lead us to hypothesize that, unlike other paramyxoviruses, HMPV could be utilizing the cellular endocytic machinery to enter the cell, and that specific residues in the HMPV F protein facilitate triggering of fusion in the low pH environment of the endosomes. To test this we will define the role of endocytosis and intracellular trafficking on HMPV viral entry using chemical inhibitors as well as dominant negative versions of key proteins involved in the initiation of various endocytic and intracellular trafficking pathways. Furthermore, we will clarify the role of HRB-linker domain interactions in HMPV F-promoted fusion and entry by introducing charged amino acid mutations in this region to determine if electrostatic repulsive forces alone are sufficient for triggering. The use of recombinant viruses carrying mutations in the HRB-linker region will also be used to assess the effect of these mutations on viral entry.
|Effective start/end date||7/1/10 → 6/30/12|
- American Heart Association Great Rivers Affiliate: $46,000.00
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