This application details a 5 year research career development program designed to explore the role of host factors in Ebola virus (EboV) entry and tropism. The program's sponsor, Dr. James Cunningham (Harvard Medical School), is a leader and has discovered receptors and host factors for both retroviral and filoviral entry. His discovery that the proteases cathepsin B &L (Cat B, CatL) are critical to entry of EboV was the first to describe the use of lysosomal proteases as host factors for enveloped virus entry. He has an excellent track record for mentoring his trainees to positions as independent investigators. My long-term goal is to become an independent physician-scientist with an expertise in emerging viral infections. As such, I have completed infectious diseases training and I am receiving research training with Dr. Cunningham. EboV causes sporadic outbreaks of a rapidly fatal hemorrhagic fever syndrome with a case mortality rate of 50-90%. Infection is characterized by cytokine storm, shock and bleeding diathesis. Studies in primates reveal that macrophages (M^) are the primary site of infection. Cat B and L are 2 of 11 members of the cysteine cathepsin (CC) protease family. These CCs have overlapping activities and, in M(j)S, cleave foreign proteins Into peptides as part of the adaptive immune response. Changes in CC expression or activity are known to modulate host THI and TH2 responses. Therefore, a detailed understanding of how CCs effect EboV entry may provide a basis to understand Mtj) tropism and the altered immune response observed during infection. Preliminary data indicates that cleavage of Ebola envelope protein (GP) is necessary but not sufficient for virus entry. Our studies indicate that infection occurs via a multi-step pathway in which CCs function in concert with other host factors. This has precedent in other Type I enveloped viruses, such as HIV, SARS and influenza, and has lead to a major effort within the Ebola community to identify these host factors;particularly toward the identification of a receptor. Despite strong evidence for the existence of an Ebola receptor, it has not been identified by use of standard techniques. We will Identify candidate host factors using a directed siRNA screen and through pull down by newly identified small molecule Ebola inhibitors. This proposal will address the following hypotheses: (1) Cleavage of GP by one or more CCs determines the tropism of filoviruses for IVI(|)S (2) There are additional host factors for filovirus entry, which contribute to tropism and pathogenesis.
(See Instructions): The viral causes of hemorrhagic fever can be rapidly fatal to humans and, while rare, are considered potential agents for bioterrorism. This research will identify the cellular determinants of how one of these viruses, EboV, enters cells. These determinants are a key component to understanding why these viruses cause hemorrhagic fever and may lead to new therapies and antivirals.
|Misasi, John; Chandran, Kartik; Yang, Jin-Yi et al. (2012) Filoviruses require endosomal cysteine proteases for entry but exhibit distinct protease preferences. J Virol 86:3284-92|
|Cote, Marceline; Misasi, John; Ren, Tao et al. (2011) Small molecule inhibitors reveal Niemann-Pick C1 is essential for Ebola virus infection. Nature 477:344-8|