Abstract
Comprehensive knowledge of the host factors required for picornavirus infection would facilitate antiviral development. Here we demonstrate roles for three human genes, TNK2, WASL, and NCK1, in infection by multiple picornaviruses. CRISPR deletion of TNK2, WASL, or NCK1 reduced encephalomyocarditis virus (EMCV), coxsackievirus B3 (CVB3), poliovirus and enterovirus D68 infection, and chemical inhibitors of TNK2 and WASL decreased EMCV infection. Reduced EMCV lethality was observed in mice lacking TNK2. TNK2, WASL, and NCK1 were important in early stages of the viral lifecycle, and genetic epistasis analysis demonstrated that the three genes function in a common pathway. Mechanistically, reduced internalization of EMCV was observed in TNK2 deficient cells demonstrating that TNK2 functions in EMCV entry. Domain analysis of WASL demonstrated that its actin nucleation activity was necessary to facilitate viral infection. Together, these data support a model wherein TNK2, WASL, and NCK1 comprise a pathway important for multiple picornaviruses.
Highlights
Picornaviruses cause a wide range of diseases including the common cold, hepatitis, myocarditis, poliomyelitis, meningitis, and encephalitis (Melnick, 1983)
We further demonstrated that Tyrosine Kinase Non-Receptor 2 (TNK2), Wiskott-Aldrich Syndrome protein Like protein (WASL), and NCK1 function in a pathway to support encephalomyocarditis virus (EMCV) virus infection with WASL and NCK1 lying downstream of TNK2
Since a role for TNK2 in clathrin-mediated endocytosis (CME) has been reported (Teo et al, 2001), we examined whether the classic CME inhibitors such as dynasore or pitstop-2 affect EMCV virus infection
Summary
Picornaviruses cause a wide range of diseases including the common cold, hepatitis, myocarditis, poliomyelitis, meningitis, and encephalitis (Melnick, 1983). Since Orsay virus is a nonenveloped, positive strand RNA virus that is evolutionarily related to the family Picornaviridae, we reasoned that the human orthologues of these genes may have a conserved role in infection by picornaviruses. Significant reductions in infection by multiple picornaviruses were observed in each of the cell lines Consistent with these in vitro data, we observed increased survival of mice lacking murine TNK2 after EMCV challenge in vivo. We further demonstrated that TNK2, WASL, and NCK1 function in a pathway to support EMCV virus infection with WASL and NCK1 lying downstream of TNK2. Loss of TNK2 led to reduced EMCV virus internalization, while both TNK2 and WASL were required for proper endocytic trafficking of EMCV These data support a model wherein TNK2, WASL, and NCK1 comprise a pathway that is important for entry by multiple picornaviruses
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