Combined Proteomics/Genomics Approach Reveals Proteomic Changes of Mature Virions as a Novel Poxvirus Adaptation Mechanism

Abstract

DNA viruses, like poxviruses, possess a highly stable genome, suggesting that adaptation of virus particles to specific cell types is not restricted to genomic changes. Cowpox viruses are zoonotic poxviruses with an extraordinarily broad host range, demonstrating their adaptive potential in vivo. To elucidate adaptation mechanisms of poxviruses, we isolated cowpox virus particles from a rat and passaged them five times in a human and a rat cell line. Subsequently, we analyzed the proteome and genome of the non-passaged virions and each passage. While the overall viral genome sequence was stable during passaging, proteomics revealed multiple changes in the virion composition. Interestingly, an increased viral fitness in human cells was observed in the presence of increased immunomodulatory protein amounts. As the only minor variant with increasing frequency during passaging was located in a viral RNA polymerase subunit and, moreover, most minor variants were found in transcription-associated genes, protein amounts were presumably regulated at transcription level. This study is the first comparative proteome analysis of virus particles before and after cell culture propagation, revealing proteomic changes as a novel poxvirus adaptation mechanism.

Estimating evolutionary changes between highly passaged and original parental lumpy skin disease virus strains

“…Four of the ORFs predicted to play a role in RNA transcription and maturation were: LW032 (a poly[A] polymerase), LW071 (an RNA polymerase subunit), LW098 (an early transcription factor) and LW099 (an intermediate transcription factor). This observation mirrors findings in cowpox viruses where the majority of the cell culture‐adapted variants occurred in the ORFs coding for the RNA polymerase subunits and transcription factors, suggesting that most of the protein regulation occurs at the transcription level rather than the genome level (Grossegesse et al., 2017). Amino acid exchanges in the palmitylated external enveloped virion envelope protein (predicted by ORF LW028), as well as the structural virion protein encoded by LW101, were observed in different high‐passaged strains.…”

Comparative Analysis of Full Genome Sequences of African Swine Fever Virus Isolates Taken from Wild Boars in Russia in 2019

“…These ORFs included EP1242L, a hybrid RNA/DNA binding RNA polymerase subunit 2; EP364R, which has a restriction endonuclease type II-like domain; O174L, the X-like DNA polymerase; NP419L, a DNA ligase; D1133L and Q706L, which both predicted that proteins belonged to the helicase super family II; and I243L, a predicted transcription factor SII ( Table 3 ). This could indicate the possibility that the principle method of acquiring genetic diversity is not on the genetic level, but rather on the transcriptomic level [ 39 ]. Alternatively, it is known that the X-type polymerase and DNA-ligase in ASFVs have low fidelity and contribute to the increased genetic drift observed within this DNA virus [ 33 , 40 ].…”

Perspective on Proteomics for Virus Detection in Clinical Samples

“…44 Additionally, it should be considered that viruses adapt in cell culture, potentially leading to proteomic changes. 45 The solution can be to identify target peptides in silico , as was recently suggested in a preprint from Orsburn and colleagues. 9 In a rigorous selection process, they identified unique tryptic peptides of SARS-CoV-2 that can potentially be used for detection.…”

“…For example, for the propagation of SARS-CoV-2, a biosafety level 3 laboratory is required according to the WHO laboratory biosafety guidance related to COVID-19 . Additionally, it should be considered that viruses adapt in cell culture, potentially leading to proteomic changes . The solution can be to identify target peptides in silico , as was recently suggested in a preprint from Orsburn and colleagues .…”