Pyk2 Autophosphorylation Mutant Inhibits HCMV-mediated SMC. (A) Schematic of Pyk2 domain structure with A457K and F402Y mutations annotated. (B) Sub-cellular localization of myc-tagged adenovirus constructs was determined via indirect immunofluorescence staining for the myc epitope. From left Ad-Pyk2 WT, Ad-Pyk2-F402Y and Ad-Pyk2-A457K in lamellipodia of adenovirus-infected PAT1 SMC. Gene expression was driven my co-infection with an adenovirus expressing the Tet-off transactivator (Ad-Trans) (C) SMC migration assays were performed on PAT1 SMC co-infected with HCMV at MOI 10 and adenoviruses expressing control Ad-Trans only or Ad-Trans+Pyk2-WT, Pyk2-A457K, or Pyk2-F402Y at MOI 50 in transwell chambers. At 2 hrs post-infection fresh medium containing 50 ng/ml CCL5 was added to the bottom chamber and cells were allowed to migrate for 48 hrs. Migrated cells in the lower chamber were counted via light microscopy. For each condition, n = 6 from two independent experiments.