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. 2005 Sep 2;6(10):961–967. doi: 10.1038/sj.embor.7400509

Figure 4.

Figure 4

Depletion of TRBP affects pre-microRNA processing in vitro but has no effect on accumulation of mature microRNAs in vivo. (A) Levels of TRBP (human immunodeficiency virus (HIV-1) transactivating response (TAR) RNA-binding protein) in 293/TRBPkd1/2 and human embryonic kidney (HEK)293T-REx control cell lines after treatment with tetracycline (Tet) for 72 h as indicated. Anti-TRBP antibody was used for western blotting. (B,C) Effect of TRBP knockdown on processing of pre-let-7 RNA in vitro (B) and on accumulation of mature microRNAs (miRNAs) in vivo (C). (B) Processing of pre-let-7 RNA was assayed in extracts from 293/TRBPkd1, 293/TRBPkd2 and 293/control-hairpin cell lines. Phosphorimage of 293/TRBPkd1 (upper panel) and quantification of data for both TRBPkd cell lines (bottom panels) are shown. (C) Total RNA isolated from TRBPkd cell lines, from two independent cultures and control cell lines was analysed by northern blot using probes specific for indicated RNAs. Quantification of data from four independent northern blots showed no significant differences in miR-16 and miR-17 levels between TRBPkd and control cultures. A representative experiment is shown. Similar results were obtained for miR-19B and let-7 miRNAs (data not shown).