A time-resolved molecular map of the macrophage response to VSV infection

Abstract

Studying the relationship between virus infection and cellular response is paradigmatic for our understanding of how perturbation changes biological systems. Immune response, in this context is a complex yet evolutionarily adapted and robust cellular change, and is experimentally amenable to molecular analysis. To visualize the full cellular response to virus infection, we performed temporal transcriptomics, proteomics, and phosphoproteomics analysis of vesicular stomatitis virus (VSV)-infected mouse macrophages. This enabled the understanding of how infection-induced changes in host gene and protein expression are coordinated with post-translational modifications by cells in time to best measure and control the infection process. The vast and complex molecular changes measured could be decomposed in a limited number of clusters within each category (transcripts, proteins, and protein phosphorylation) each with own kinetic parameter and characteristic pathways/processes, suggesting multiple regulatory options in the overall sensing and homeostatic program. Altogether, the data underscored a prevalent executive function to phosphorylation. Resolution of the molecular events affecting the RIG-I pathway, central to viral recognition, reveals that phosphorylation of the key innate immunity adaptor mitochondrial antiviral-signaling protein (MAVS) on S328/S330 is necessary for activation of type-I interferon and nuclear factor κ B (NFκB) pathways. To further understand the hierarchical relationships, we analyzed kinase–substrate relationships and found RAF1 and, to a lesser extent, ARAF to be inhibiting VSV replication and necessary for NFκB activation, and AKT2, but not AKT1, to be supporting VSV replication. Integrated analysis using the omics data revealed co-regulation of transmembrane transporters including SLC7A11, which was subsequently validated as a host factor in the VSV replication. The data sets are predicted to greatly empower future studies on the functional organization of the response of macrophages to viral challenges.

Time-resolved proteomics of adenovirus infected cells

“…In addition, only one of the most relevant canonical pathways was found common when RNA and protein data were compared. Moreover, in a recent publication where the expression dynamics of transcripts, proteins and phosphoproteins upon VSV infection of macrophages was studied, surprising differences between the three levels were demonstrated [ 17 ].…”

“…time series, to describe these processes [ 13 , 14 , 15 ]. Time series have been previously used to study the Herpes Simplex Virus Type 1 infection of human foreskin fibroblast [ 16 ], the macrophage response to Vesicular Stomatitis Virus infection [ 17 ], or the infection by H9N2 influenza virus in a human gastric carcinoma cell line [ 18 ], but also to investigate essential cellular functions [ 19 ]. Among the different strategies developed for the comparison of protein changes [ 20 ], the incorporation of stable isotopes using metabolic labelling of amino acids in cell culture (SILAC) has demonstrated to decrease the errors in quantification, and it offers good precision when comparing biological samples [ 21 ].…”

Coactivation of TLR2 and TLR8 in Primary Human Monocytes Triggers a Distinct Inflammatory Signaling Response

“…Post-translational modifications (PTMs) plays a crucial role in the activation and regulation of this complex response ( Liu et al, 2016 ). PTMs such as phosphorylation have distinct patterns depending on the type of activation ( Kandasamy et al, 2016 ).…”

TRAF3IP3 negatively regulates cytosolic RNA induced anti-viral signaling by promoting TBK1 K48 ubiquitination

“…4h, i). Similarly, profiling of Traf3ip3 in RAW264.7 cells using Infectome Map 38 showed Traf3ip3 also appeared stable after VSV challenge ( Supplementary Fig. 4j).…”