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. 2004 Feb 15;18(4):423–434. doi: 10.1101/gad.285504

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Copyright © 2004, Cold Spring Harbor Laboratory Press

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Figure 6. — Phosphorylation of S199 impairs the interaction of TIF-IA with Pol I. (A) Rapamycin treatment inhibits the interaction between TIF-IA and Pol I. Lysates from mock- or rapamycin-treated HeLa cells expressing Flag-tagged TIF-IA were incubated with anti-Flag antibodies, and immunoprecipitated TIF-IA and Pol I were visualized on Western blots using anti-Flag, anti-RPA116, or anti-PAF67 antibodies. (B) A mutant that mimics phosphorylation of TIF-IA at S199 fails to interact with Pol I and TIF-IB. Flag-tagged TIF-IA or the indicated point mutants were overexpressed in HEK293T cells and immunoprecipitated with anti-Flag antibodies. Coprecipitated Pol I and TIF-IB/SL1 were visualized on Western blots with anti-RPA116 and anti-TAF_I110 antibodies.