Skip to main content
. 2003 Mar;23(5):1688–1702. doi: 10.1128/MCB.23.5.1688-1702.2003

FIG. 6.

FIG. 6.

Effect of granulin binding on kinase activity of P-TEFb. (A) GST-GrnCDE pulls down P-TEFb from 293T cell WCE. Immunoblot (IB) analysis was performed with anti-CDK9 antibody. CDK9 was detected in complexes bound to GST-GrnCDE (lane 4) and GST-Tat48Δ (lane 3) but not GST alone (lane 2). WCE was used as control for the presence of CDK9 (lane 1). (B) Complexes bound to GST-GrnCDE and GST-Tat48Δ were tested in kinase assays in the absence and presence of CTD4 peptide. CTD4 was phosphorylated by GST-Tat48Δ-bound complexes (lane 5) but not by GST-GrnCDE-bound complexes (lane 4). GST-GrnCDE was phosphorylated in the absence or presence of CTD4 (lanes 3 and 4). Neither GST-GrnCDE nor CTD4 was phosphorylated in the absence of WCE (lanes 1 and 2). (C) CDK9 immunocomplexes from 293T WCE phosphorylated purified GST-GrnCDE (lane 1) or GST-CTD (lane 5). The phosphorylation was inhibited by 10 μM DRB (lanes 2 and 6). Increasing concentration of His peptide 511 to 530 inhibited the phosphorylation of GST-GrnCDE but not of GST-CTD.