Research Article: Adenovirus early region 3 RIDα protein limits NFκB signaling through stress-activated EGF receptors

Date Published: August 19, 2019

Publisher: Public Library of Science

Author(s): Xuehuo Zeng, Cathleen R. Carlin, Patrick Hearing.

http://doi.org/10.1371/journal.ppat.1008017

Abstract

The host limits adenovirus infections by mobilizing immune systems directed against infected cells that also represent major barriers to clinical use of adenoviral vectors. Adenovirus early transcription units encode a number of products capable of thwarting antiviral immune responses by co-opting host cell pathways. Although the EGF receptor (EGFR) was a known target for the early region 3 (E3) RIDα protein encoded by nonpathogenic group C adenoviruses, the functional role of this host-pathogen interaction was unknown. Here we report that incoming viral particles triggered a robust, stress-induced pathway of EGFR trafficking and signaling prior to viral gene expression in epithelial target cells. EGFRs activated by stress of adenoviral infection regulated signaling by the NFκB family of transcription factors, which is known to have a critical role in the host innate immune response to infectious adenoviruses and adenovirus vectors. We found that the NFκB p65 subunit was phosphorylated at Thr254, shown previously by other investigators to be associated with enhanced nuclear stability and gene transcription, by a mechanism that was attributable to ligand-independent EGFR tyrosine kinase activity. Our results indicated that the adenoviral RIDα protein terminated this pathway by co-opting the host adaptor protein Alix required for sorting stress-exposed EGFRs in multivesicular endosomes, and promoting endosome-lysosome fusion independent of the small GTPase Rab7, in infected cells. Furthermore RIDα expression was sufficient to down-regulate the same EGFR/NFκB signaling axis in a previously characterized stress-activated EGFR trafficking pathway induced by treatment with the pro-inflammatory cytokine TNF-α. We also found that cell stress activated additional EGFR signaling cascades through the Gab1 adaptor protein that may have unappreciated roles in the adenoviral life cycle. Similar to other E3 proteins, RIDα is not conserved in adenovirus serotypes associated with potentially severe disease, suggesting stress-activated EGFR signaling may contribute to adenovirus virulence.

Partial Text

Human adenoviruses provide an excellent example of how viruses adapt host cell machinery to invade cells, gain access to the nucleus to replicate, assemble new viral particles, and spread in the host [1,2,3]. The host limits adenovirus infections by mobilizing innate immune systems that activate inflammatory or cytotoxic responses directed against infected cells [3,4,5,6]. These host defense mechanisms also represent a major barrier to the use of adenovirus vectors with many important clinical applications, ranging from cancer gene therapy to vaccine development [3,4,5,6]. In addition to specialized immune cells that secrete pro-inflammatory cytokines at sites of infection, immune and non-immune target cells both rely on cell autonomous innate immunity to defend against the immediate threat of infection [7]. Adenovirus circumvents various innate defense mechanisms by virtue of viral proteins encoded by early transcription units that strike a balance between the elimination of virus and immune-mediated tissue injury [8]. The study of cellular pathways used by viruses has led to many significant advances in eukaryotic cell and molecular biology [9]. Adenovirus early region 3 (E3) gene products in particular have been powerful tools for discovering new mechanisms in the fields of intracellular protein and lipid trafficking [10,11,12].

Our studies have revealed three major new findings. First, incoming viral particles induced a non-canonical pathway of stress-activated EGFR trafficking and signaling prior to nuclear translocation and transcription of viral DNA in epithelial target cells. Second, stress-induced EGFR signaling was required for a site-specific phosphorylation event with a pivotal role in NFκB-p65 function. Third, the adenoviral RIDα protein attenuated the EGFR/NFκB signaling axis in the context of an acute adenovirus infection, and as an independently expressed transgene in cells stimulated with the pro-inflammatory cytokine TNF-α. The following working model interprets data presented here in the context of the current literature (see Fig 9 for a schematic diagram).

 

Source:

http://doi.org/10.1371/journal.ppat.1008017

 

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