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. Author manuscript; available in PMC: 2019 Jul 31.
Published in final edited form as: Cell Rep. 2019 Jul 9;28(2):449–459.e5. doi: 10.1016/j.celrep.2019.04.020

Figure 2. SAMHD1 Is Phosphorylated by EBV Protein Kinase BGLF4.

Figure 2.

(A) SAMHD1 is phosphorylated upon lytic induction of EBV. Western blot analysis was performed on cell lysates from Akata-BX1 (EBV+) and Akata 4E3 (EBV−) cells using antibodies as indicated. The cells were either untreated (0 hr) or treated with anti-human IgG for 24, 48, or 72 h to induce lytic reactivation. Arrowhead denotes the major 72-kDa phospho-SAMHD1 band, and asterisk denotes a 69-kDa non-specific band or a phospho-SAMHD1 band derived from a SAMHD1 isoform protein.

(B) SAMHD1 is phosphorylated in EBV-replicating HeLa cells. Western blot analysis was performed on cell lysates from HeLa (EBV+) transfected with EBV ZTA, RTA, or ZTA plus RTA as indicated for 48 h to induce lytic reactivation. The phosphorylation of SAMHD1 correlated with BGLF4 expression.

(C) SAMHD1 is phosphorylated by EBV protein kinase BGLF4 in vitro. Recombinant SAMHD1 protein was mixed with purified wild-type or KD BGLF4 for 30 min at 30°C. As a positive control, SAMHD1 was incubated with CDK1/Cyclin B (lane 1), which is a known kinase for SAMHD1. As negative controls, either kinase or ATP was omitted in the reaction mixture (lanes 2, 5, and 6). The phospo-SAMHD1, SAMHD1, and BGLF4 were detected by western blot using antibodies as indicated.

See also Figure S1.