Host Src controls gallid alpha herpesvirus 1 intercellular spread in a cellular fatty acid metabolism-dependent manner

Abstract

A B S T R A C TViral spread is considered a promising target for antiviral therapeutics, but the associated mechanisms remain unclear for gallid alpha herpesvirus 1 (ILTV). We previously identified proto-oncogene tyrosine-protein kinase Src (Src) as a crucial host determinant of ILTV infection. The present study revealed accelerated spread of ILTV upon Src inhibition. This phenomenon was independent of either viral replication or the proliferation of infected cells and could not be compromised by neutralizing antibody. Neither extracellular vesicles nor the direct cytosol-to-cytosol connections between adjacent cells contributed to the enhanced spread of ILTV upon Src inhibition. Further genome-wide transcriptional profile analyses in combination with functional validation identified fatty acid metabolism as an essential molecular event during modulation of the intercellular spread and subsequent cytopathic effect of ILTV by Src. Overall, these data suggest that Src controls the cell-to-cell spread of ILTV in a cellular fatty acid metabolism-dependent manner, which determines the virus's cytopathic effect.

Global exploration of the metabolic requirements of gallid alphaherpesvirus 1

“…To determine whether the metabolic requirements of ILTV infection revealed in chicken cell lines also exist in primary cells, further validations were performed in primary cells isolated from specific pathogen free (SPF) chicken embryos, including chicken embryo kidney cells (CEK) and chicken embryo liver cells (CEL). An ILTV-LSJ09 strain expressing enhanced green fluorescent protein (EGFP), which exhibits similar viral replication kinetics and CPEs of infection as the wild type stain [13,15], was employed to monitor viral infection in CEK and CEL. Both CEK and CEL comprised a heterogeneous population of cells, which could be divided morphologically into fibroblast-like cells and non-fibroblast like cells ( S4 Fig and Fig 8B).…”

“…This strain can be propagated in a chemically immortalized leghorn male hepatoma (LMH) cell line with clear CPEs observed [27,28]. An ILTV-LSJ09 strain expressing enhanced green fluorescent protein (EGFP), which exhibits no significant difference from the wild type in either viral replication or the CPEs of infection as described previously [13,15], is stored at the Harbin Veterinary Research Institute of CAAS. The chicken fibroblast cell line DF-1 (ATCC CRL-12203) and the chicken hepatoma cell line LMH (ATCC CRL-2117) were maintained in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum (FBS), 100 units/ml penicillin, 100 μg/ml streptomycin and 2 mM L-glutamine.…”

“…During this process, many metabolic signaling pathways are altered in host cells, which might play an important role in ILTV replication. At minimum, fatty acid metabolism has been found to be essential for the cell-to-cell transmission of ILTV [ 13 ]. Therefore, elucidation of the metabolic requirements of ILTV infection may increase current understanding of herpesvirus-host metabolic interplay and inspire further studies in this direction.…”

“…Although primary cells can better mimic the in vivo process of virus infection, the complexity, heterogeneity, and short lifespan of primary cells during in vitro culture limit their use in mechanistic studies. Thus, LMH cells have been commonly used for the investigation of ILTV infection and proven to be a reliable in vitro model [ 12 , 13 , 15 – 18 ]. However, despite being capable of exploring the metabolic characteristics of ILTV, this model may not be sensitive enough to investigate metabolic shift at the early stage of virus infection, such as the initial stage of viral transcription, due to that tumor cells often have distinct metabolic patterns to meet the rapid energy requirements [ 19 , 20 ].…”

Fos Facilitates Gallid Alpha-Herpesvirus 1 Infection by Transcriptional Control of Host Metabolic Genes and Viral Immediate Early Gene

“…Western blotting was performed strictly according to previously described procedures [19]. Briefly, cells were washed with ice-cold PBS and soluble proteins were extracted with cell lysis buffer (100 mM Tris-HCl (pH = 8), 150 mM NaCl, 1% NP-40, phosphatase and protease inhibitor cocktail tablets (Abcam, Shanghai, China)) according to the manufacturer's protocol.…”