Figure 2.

CIA1–CIA2B–MMS19 targeting complex interacts with the C terminus and CIA2A with the N terminus of viperin. A, FLP-IN T Rex cells were treated with tetracycline (1 μg/ml) to induce expression either of wild-type viperin (Wt), viperin versions lacking the TN50 or TC20 residues, or the viperin Fe/S cluster binding mutant M1 (Cys to Ala mutations). After cell growth for 1 day, viperin was immunoprecipitated, and both whole cell lysates (Input) and immunoprecipitates (IP-Viperin) were analyzed by immunoblotting. Tubulin served as loading control. B, HEK293T cells were transiently transfected with plasmids encoding N-terminally FLAG-tagged wild-type viperin, viperin mutants TN50 and TC20, and viperin fragments consisting of the C100 or C70 residues. The FLAG proteins were precipitated by an anti-FLAG antibody, and samples were analyzed by immunoblotting. C, HEK293T cells were transiently transfected as in B and additionally received a plasmid encoding Myc-tagged CIA2A. Anti-FLAG–viperin IP and sample analysis were performed as in B.