The long term goal of this research is to understand the mechanism by which enteroviruses such as poliovirus (PV) and Coxsackievirus (CVB3) inactivate translation of nearly all cellular mRNA while stimulating efficient translation of viral mRNA in infected cells. We have shown that cleavage of the translation initiation factor eIF4G will block assembly of new capped mRNA on ribosomes, and that cleavage of PABP is required in concert with eIF4G to completely block translation. PABP cleavage affects late steps in translation that are presently undefined, however likely interrupt ribosome recycling via 5'-3'interactions on mRNA. The mechanism of ribosome recycling and its biochemical requirements are unknown. In addition, enteroviruses and probably most other plus strand RNA viruses must abruptly shut down translation of the infecting viral genome before viral RNA synthesis can begin. We hypothesize that PABP cleavage is also required for inhibition of viral translation. Translation regulation mechanisms involving PABP are now thought to interface with mRNA decay mechanisms on several levels. In addition, microRNAs, which bind to 3'UTR of targeted cellular mRNAs, also silence translation by unknown mechanisms that somehow result in transit of mRNAs from polysomes to other cell compartments called P-bodies and stress granules. We have discovered that G3BP, a key factor that nucleates formation of stress granules, is cleaved in PV-infected cells by 3C protease. Thus, the virus is attacking the overall translation regulatory apparatus at a new level.
The aims i n this proposal will determine the role of PABP cleavage in the switch from viral translation to RNA replication, will determine the role of ribosome recycling in this switch, and will investigate the function of G3BP cleavage on the viral replication cycle and microRNA-translation silencing. These results will provide new fundamental information on how cells regulate gene expression at the translation level, and the interplay between translation initiation factors and mRNA silencing/decay pathways that will be useful in studies of viral regulation of gene expression and cellular stress responses in cancer and apoptosis. This work investigates new ways that poliovirus, the prototype human picornavirus, interferes with host translation and regulation of gene expression by stress granules and microRNAs. These mechanisms are fundamental for cell gene regulation and are disrupted in cancer and numerous non-viral diseases. These experiments will give us more insight how a large class of human pathogens invade and kill cells (cytopathology) and potentially provide new avenues for disease prevention and we will learn more about gene regulation in human cells.
|Reineke, Lucas C; Tsai, Wei-Chih; Jain, Antrix et al. (2017) Casein Kinase 2 Is Linked to Stress Granule Dynamics through Phosphorylation of the Stress Granule Nucleating Protein G3BP1. Mol Cell Biol 37:|
|Tsai, Wei-Chih; Reineke, Lucas C; Jain, Antrix et al. (2017) Histone arginine demethylase JMJD6 is linked to stress granule assembly through demethylation of the stress granule nucleating protein G3BP1. J Biol Chem :|
|Tsai, Wei-Chih; Gayatri, Sitaram; Reineke, Lucas C et al. (2016) Arginine Demethylation of G3BP1 Promotes Stress Granule Assembly. J Biol Chem 291:22671-22685|
|Lloyd, Richard E (2016) Enterovirus Control of Translation and RNA Granule Stress Responses. Viruses 8:93|
|Reineke, Lucas C; Lloyd, Richard E (2015) The stress granule protein G3BP1 recruits protein kinase R to promote multiple innate immune antiviral responses. J Virol 89:2575-89|
|Reineke, Lucas C; Kedersha, Nancy; Langereis, Martijn A et al. (2015) Stress granules regulate double-stranded RNA-dependent protein kinase activation through a complex containing G3BP1 and Caprin1. MBio 6:e02486|
|Lloyd, Richard E (2015) Nuclear proteins hijacked by mammalian cytoplasmic plus strand RNA viruses. Virology 479-480:457-74|
|Dougherty, Jonathan D; Tsai, Wei-Chih; Lloyd, Richard E (2015) Multiple Poliovirus Proteins Repress Cytoplasmic RNA Granules. Viruses 7:6127-40|
|Dougherty, Jonathan D; Reineke, Lucas C; Lloyd, Richard E (2014) mRNA decapping enzyme 1a (Dcp1a)-induced translational arrest through protein kinase R (PKR) activation requires the N-terminal enabled vasodilator-stimulated protein homology 1 (EVH1) domain. J Biol Chem 289:3936-49|
|Feng, Qian; Langereis, Martijn A; Lork, Marie et al. (2014) Enterovirus 2Apro targets MDA5 and MAVS in infected cells. J Virol 88:3369-78|
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