We estimated activity regulation for 97 kinases based on the regulation of their known substrates (Ochoa et al., 2016; Hernandez-Armenta et al., 2017; Table S4), with the strongest regulation linked to viral entry (0–2 h) and late replication/egress (24 h). The kinases predicted to be most strongly activated (Figures 4B and S2A) include several members of the p38 pathway, including p38ɣ (MAPK12), CK2 (CSNK2A1/2), Ca2+/calmodulin-dependent protein kinase (CAMK2G), and the guanosine monophosphate (GMP)-dependent protein kinases PRKG1/2, which can inhibit Rho signaling. Kinases predicted to be downregulated include several cell cycle kinases (CDK1/2/5 and AURKA), cell growth-related signaling pathway kinases (PRKACA, AKT1/2, MAPK1/3, and PIM1), and the cytoskeleton regulators (PAK1), among others. Kinase activity estimates based on the 24-h mock control gave highly correlated results (r = 0.81), identifying the same set of highly regulated kinases (Figure S2D). Some of the changes in kinase activity can be directly linked to host-viral protein interactions (Figure 4C). Among the 10 interacting kinases detected in a virus-host protein-protein interaction map (Gordon et al., 2020), an increase in activity for CK2 and a decrease for MARK2 and PRKACA were observed (Figure 4C). Of note, although we predict decreased activity for PRKACA based on phosphorylation of its substrates, we simultaneously detected a significant increase in T198 phosphorylation (8, 12, and 24 h post infection) within its activation loop, suggesting an increase in PRKACA activity. It is possible that Nsp13 is sequestering active PRKACA away from its typical substrates.
Figure S2 Full Kinase Activities, Correlated Conditions, and Regulated Complexes, Related to Figure 4
(A) Changes in predicted kinase activities across different time points post-infection. (B) Correlation of kinase activity profiles of each time point with other biological conditions. Kinase activities were estimated for a wide-range of biological conditions obtained from previously published phosphoproteomics datasets (Ochoa et al., 2016). (C) Changes in phosphorylation in protein complexes. Overall phosphorylation change (-log10 (p value) of a protein complex was derived from change in phosphorylation of sites in member proteins. (D) Kinase activity estimates when using either the 0- or 24-h mock controls for those top regulated kinase activities from the 0-h control comparison.