Host Cellular Protein TRAPPC6AΔ Interacts with Influenza A Virus M2 Protein and Regulates Viral Propagation by Modulating M2 Trafficking.

Pengyang Zhu,Libin Liang,Nan Sun,Junping Li,Yuan Zhou,Guangwen Wang,Li Jiang,Qingqing Zhao,Jie Zhang,Yuhui Zhao,Hualan Chen,Shuitao Jiang,Chengjun Li,Jinguang Wang,Weiyu Luo,Xinyuan Shao,Terence S Dermody

doi:10.1128/jvi.01757-16

Pengyang Zhu, Libin Liang + Show 15 more

Open Access

https://doi.org/10.1128/jvi.01757-16

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Abstract

ABSTRACTInfluenza A virus (IAV) matrix protein 2 (M2) plays multiple roles in the early and late phases of viral infection. Once synthesized, M2 is translocated to the endoplasmic reticulum (ER), travels to the Golgi apparatus, and is sorted at the trans-Golgi network (TGN) for transport to the apical plasma membrane, where it functions in virus budding. We hypothesized that M2 trafficking along with its secretory pathway must be finely regulated, and host factors could be involved in this process. However, no studies examining the role of host factors in M2 posttranslational transport have been reported. Here, we used a yeast two-hybrid (Y2H) system to screen for host proteins that interact with the M2 protein and identified transport protein particle complex 6A (TRAPPC6A) as a potential binding partner. We found that both TRAPPC6A and its N-terminal internal-deletion isoform, TRAPPC6A delta (TRAPPC6AΔ), interact with M2. Truncation and mutation analyses showed that the highly conserved leucine residue at position 96 of M2 is critical for mediating this interaction. The role of TRAPPC6AΔ in the viral life cycle was investigated by the knockdown of endogenous TRAPPC6AΔ with small interfering RNA (siRNA) and by generating a recombinant virus that was unable to interact with TRAPPC6A/TRAPPC6AΔ. The results indicated that TRAPPC6AΔ, through its interaction with M2, slows M2 trafficking to the apical plasma membrane, favors viral replication in vitro, and positively modulates virus virulence in mice.IMPORTANCE The influenza A virus M2 protein regulates the trafficking of not only other proteins but also itself along the secretory pathway. However, the host factors involved in the regulation of the posttranslational transport of M2 are largely unknown. In this study, we identified TRAPPC6A and its N-terminal internal-deletion isoform, TRAPPC6AΔ, as interacting partners of M2. We found that the leucine (L) residue at position 96 of M2 is critical for mediating this interaction, which leads us to propose that the high level of conservation of 96L is a consequence of M2 adaptation to its interacting host factor TRAPPC6A/TRAPPC6AΔ. Importantly, we discovered that TRAPPC6AΔ can positively regulate viral replication in vitro by modulating M2 trafficking to the plasma membrane.

Highlights

Influenza A virus (IAV) matrix protein 2 (M2) plays multiple roles in the early and late phases of viral infection
We found that the leucine (L) residue at position 96 of M2 is critical for mediating this interaction, which leads us to propose that the high level of conservation of 96L is a consequence of M2 adaptation to its interacting host factor TRAPPC6A/TRAPPC6AΔ
TRAPPC6A⌬ positively modulates influenza virus infection. We demonstrated that both TRAPPC6A and TRAPPC6AΔ interact with M2 of IAV

Summary

Introduction

Influenza A virus (IAV) matrix protein 2 (M2) plays multiple roles in the early and late phases of viral infection. We used a yeast two-hybrid (Y2H) system to screen for host proteins that interact with the M2 protein and identified transport protein particle complex 6A (TRAPPC6A) as a potential binding partner We found that both TRAPPC6A and its N-terminal internal-deletion isoform, TRAPPC6A delta (TRAPPC6AΔ), interact with M2. The proton channel activity of M2 can raise the pH of the trans-Golgi network (TGN) to prevent the premature low-pH conformational change of viral hemagglutinin (HA) during its transport to the cell surface This function is important for highly pathogenic H7 influenza viruses, whose HA can be cleaved by ubiquitous proteases and is more susceptible to low-pH-induced premature maturation [18, 19] and essential to maintain 2009 pandemic H1N1 virus HA fusion competence during transport to the cell surface [20]. The M2 proton channel activity can activate the NLRP3 inflammasome of macrophages and dendritic cells [21]

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