Fig 5.

orf36 modulates global HDAC activity throughout lytic infection of primary macrophages. (A to C) Primary macrophages were mock infected or infected at an MOI of 1 or 10 with wild-type or orf36 mutant MHV68 viruses. HDAC enzymatic activity, total protein concentration, and HDAC1, HDAC2, and β-actin protein levels were measured in whole-cell lysates collected at the indicated times postinfection. HDAC activity was normalized to total protein content and, subsequently, to mock-infected lysates. In parallel, HDAC activity was measured in cell lysates spiked with TSA. HDAC activity data were pooled from 3 to 6 independent experiments; each condition was assessed in two replicates within each experiment. Densitometric analysis was used to measure HDAC1 and -2 protein content, normalized to β-actin. Normalized values are listed below the respective lanes. (D) Primary macrophages were nucleofected with the indicated plasmids, and HDAC activity and protein levels were measured as above at 6 h postnucleofection. (E) Primary macrophages were mock infected or infected with 36F MHV68 at an MOI of 10. orf36 and β-actin protein levels were measured at the indicated times postinfection. All error bars represent one standard error of the mean. *, P < 0.05; ***, P < 0.001.