The Ras–PI3K Signaling Pathway Is Involved in Clathrin-Independent Endocytosis and the Internalization of Influenza Viruses

Yoichiro Fujioka,Yusuke Ohba,Tomoe Hattori,Takehiko Sasaki,Junko Sasaki,Tadaaki Miyazaki,Masumi Tsuda

doi:10.1371/journal.pone.0016324

Abstract

BackgroundInfluenza virus infection causes highly contagious, severe respiratory disorders and gives rise to thousands of deaths every year; however, the efficacy of currently approved defense strategies, including vaccines and neuraminidase inhibitors, is limited because the virus frequently acquires resistance via antigen drift and reassortment. It is therefore important to establish a novel, effective therapeutic strategy that is effective irrespective of viral subtype.Methodology/Principal FindingsHere, we identify the Ras–phosphoinositide 3-kinase (PI3K) signaling pathway as a host-cell regulatory mechanism for influenza virus entry. The binding of Ras to PI3K is specifically involved in clathrin-independent endocytosis, endosomal maturation, and intracellular transport of viruses, which result in decreased infectious efficacy of different subtypes of influenza viruses in cells lacking the Ras–PI3K interaction. Moreover, influenza virus infection indeed triggered Ras activation and subsequent PI3K activation in early endosomes.Conclusions/SignificanceTaken together, these results demonstrate that the Ras–PI3K signaling axis acts as a host-oriented mechanism for viral internalization. Given that virus incorporation is a process conserved among virus subtypes and species, this signaling pathway may provide a target for potent, well-tolerated prophylactics and therapeutics against a broad range of viruses.

Highlights

Influenza spreads around the world in seasonal epidemics, resulting in the death of hundreds of thousands of people annually—millions in pandemic years
In order to test whether phosphoinositide-3 kinases (PI3Ks) participates in the regulation of endocytosis, we first prepared mouse embryonic fibroblasts (MEFs) from wild-type mice and mice deficient in Pik3cg
The treatment of wild-type MEFs with the pan-PI3K inhibitor LY294002 significantly reduced the uptake of dextran, but not that of transferrin (Figure 1A, B), confirming that PI3K is involved in clathrin-independent endocytosis, as reported previously [20]

Summary

Introduction

Influenza spreads around the world in seasonal epidemics, resulting in the death of hundreds of thousands of people annually—millions in pandemic years. Neuraminidase (NA) inhibitors such as oseltamivir and zanamivir have been efficacious against the current H1N1 strain of swine origin; the virus may acquire resistance to the available antiviral drugs. Vaccines are available for the prevention and control of influenza virus infection, they need to be frequently revised—typically every 1–3 years in the case of seasonal influenza vaccines—to accommodate mutations in the HA and NA proteins of the circulating viruses (antigen drift). Influenza virus infection causes highly contagious, severe respiratory disorders and gives rise to thousands of deaths every year; the efficacy of currently approved defense strategies, including vaccines and neuraminidase inhibitors, is limited because the virus frequently acquires resistance via antigen drift and reassortment. It is important to establish a novel, effective therapeutic strategy that is effective irrespective of viral subtype

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Results

Conclusion