title: Chemoproteomic profiling identifies changes in DNA-PK as markers of early dengue virus infection.
abstract: Many cellular factors are regulated via mechanisms affecting protein conformation, localization, and function that may be undetected by most commonly used RNA- and protein-based profiling methods that monitor steady-state gene expression. Mass-spectrometry-based chemoproteomic profiling provides alternatives for interrogating changes in the functional properties of proteins that occur in response to biological stimuli, such as viral infection. Taking dengue virus 2 (DV2) infection as a model system, we utilized reactive ATP- and ADP-acyl phosphates as chemical proteomic probes to detect changes in host kinase function that occur within the first hour of infection. The DNA-dependent protein kinase (DNA-PK) was discovered as a host enzyme with significantly elevated probe labeling within 60 min of DV2 infection. Increased probe labeling was associated with increased DNA-PK activity in nuclear lysates and localization of DNA-PK in nucleoli. These effects on DNA-PK were found to require a postfusion step of DV2 entry and were recapitulated by transfection of cells with RNA corresponding to stem loop B of the DV2 5' untranslated region. Upon investigation of the potential downstream consequences of these phenomena, we detected a modest but significant reduction in the interferon response induced by DV2 in cells partially depleted of the Ku80 subunit of DNA-PK. These findings identify changes in DNA-PK localization and activity as very early markers of DV2 infection. More broadly, these results highlight the utility of chemoproteomic profiling as a tool to detect changes in protein function associated with different cell states and that may occur on very short time scales.
authors.affiliation: Department of Microbiology and Immunobiology, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Michael L
authors.initials: ML
authors.lastname: Vetter
authors.forename: Mary A
authors.initials: MA
authors.lastname: Rodgers
authors.forename: Matthew P
authors.initials: MP
authors.lastname: Patricelli
authors.forename: Priscilla L
authors.initials: PL
authors.lastname: Yang
mesh_terms: D020121:5' Untranslated Regions; D015415:Biomarkers / Q000378:metabolism*; D015153:Blotting, Western; D002460:Cell Line; D051747:DNA-Activated Protein Kinase / Q000378:metabolism*; D003715:Dengue / Q000201:enzymology*; D006801:Humans; D040901:Proteomics; D015398:Signal Transduction
chemical_list: D020121:5' Untranslated Regions; D015415:Biomarkers; D051747:DNA-Activated Protein Kinase