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. Author manuscript; available in PMC: 2013 May 11.
Published in final edited form as: Mol Cell. 2012 Apr 5;46(3):325–334. doi: 10.1016/j.molcel.2012.03.007

Figure 2. Siah1 depletion suppresses ELL2 polyubiquitination and promotes ELL2 stability, SEC formation and SEC-dependent HIV-1 transcription.

Figure 2

A. The HA-Siah1 expression plasmid was co-transfected into HeLa cells with an empty vector (ctrl) or plasmids expressing the indicated shRNAs targeting Siah1. HA-Siah1 and α-tubulin present in cell lysates were examined by Western blotting. B. Whole cell lysates (WCE) of cells expressing the indicated shSiah1 sequences were analyzed by Western Blotting (WB) for the indicated proteins. C. WCE examined in B were subjected to immunoprecipitation with either anti-CDK9 Ab or total rabbit (ctl.) IgG and immunoprecipitates were analyzed by WB for the indicated proteins. D. Polyubiquitinated ELL2-F(Ub)n isolated by pull-down with Ni2+-NTA beads and total ELL2-F in WCE of cells expressing the indicated shSiah1 sequences were examined by anti-Flag WB. E. Luciferase activities were measured in extracts of cells co-transfected with the indicated shSiah1-expressing constructs, the HIV-1 LTR-luciferase reporter gene, and a vector expressing Tat or nothing. The activity in cells expressing the non-effective shSiah1 #3 but not Tat was set to 1. The error bars represent mean +/− SD from three independent sets of transfection.