Proteotype profiling unmasks a viral signalling network essential for poxvirus assembly and transcriptional...

Area of Science:

• Virology
• Molecular Biology
• Biochemistry

Background:

• Poxviruses utilize dual-specificity enzymes, F10 kinase and H1 phosphatase, for context-dependent signaling.
• The precise substrate targets and system-level functions of these enzymes are not fully understood.

Purpose of the Study:

• To systematically define the viral signaling network regulated by F10 kinase and H1 phosphatase.
• To elucidate the genotype-phenotype relationships governed by these viral enzymes.

Main Methods:

• Utilized phosphoproteomic screening of cells infected with wild-type and mutant vaccinia viruses (F10 and H1).
• Performed quantitative cross-comparison of phosphosites to identify viral proteins regulated by F10 and H1.
• Integrated proteotype data with genotype-phenotype analysis.

Main Results:

• Identified 33 F10 and/or H1 phosphosites across 17 viral proteins.
• Discovered a hidden hypercleavage phenotype in H1-deficient virions due to I7 protease phosphorylation.
• Uncovered that phosphorylation of the early transcription factor A7 underlies the transcription-deficient phenotype in H1 mutants.

Conclusions:

• Quantitative proteomic screens combined with mutant viruses effectively reveal masked proteotype-phenotype-genotype relationships.
• This approach provides novel insights into poxvirus signaling pathways and enzyme functions.