Profiling whole-tissue metabolic reprogramming during cutaneous poxvirus infection and clearance

Background Vaccinia virus (VACV) infection induces prominent changes in host cell metabolism. Little is known about the global metabolic reprogramming that takes place in the whole tissue during viral infection. Here, we performed an unbiased longitudinal metabolomics study in VACV-infected mice to investigate metabolic changes in the tissue during infection. We assessed metabolites in homogenized skin over time in the presence or absence of antigen-specific T cells using untargeted mass spectrometry. VACV infection induced several significant metabolic changes, including in the levels of nucleic acid metabolites (reflecting the impact of viral replication on the skin metabolome). Furthermore, monocyte- and antiviral T cell-produced metabolites, including itaconic acid, glutamine, and glutathione, were significantly increased following infection, highlighting the immune response’s contribution to the global skin metabolome. Additional RNA-Seq of infected skin tissue recapitulated transcriptional changes identified via metabolomics. Overall, our study reveals the metabolic balance of viral replication and the antiviral immune response in the skin and identifies metabolic pathways that could contribute to cutaneous poxvirus control in vivo. Ears were collected on indicated day post-infection (dpi) and immediately placed in buffer RLT (Qiagen). Tissues were homogenized, and RNA was isolated using Lysing Matrix S (1/8”) metal beads (MPBiomedicals) and a FastPrep-24 Instrument (MPBiomedicals). RNA was then purified using a Qiagen RNAEasy Mini Kit (Qiagen) according to the manufacturer’s protocol. An on-column DNAse digestion was performed prior to RNA elution. RNA concentration and purity were accessed using an Agilent TapeStation 4200. Samples were sequenced on a NovoSeq 6000 S1 using Illumina® Stranded Total RNA Prep, Ligation with Ribo-Zero Plus and paired-end sequencing.