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. 2003 Nov 15;17(22):2747–2752. doi: 10.1101/gad.1140303

Figure 3.

Figure 3.

Effects of altered linker length between the P1b helix and the template on template boundary definition. (A) Schematic diagram of the mutant hTR pseudoknot fragments. The U-residue insertions made in the pseudoknot mutants, 32–195(+1U) and 32–195(+2U), are indicated by open triangles. (B) Telomerase activity assay of in vitro reconstituted telomerase containing the linker-length mutant RNAs. Various pseudoknot RNA fragments— hTR32–195-wt, hTR32–195(+1U), and hTR32–195(+2U)—were assembled with hTR–CR4–CR5 RNA and in vitro expressed hTERTHA protein. In vitro reconstituted enzyme was immunopurified and assayed for activity. The activity assay was carried out in either the absence or the presence of 0.5 mM dCTP. Bands representing incorporation past the normal template are marked with white triangles. (C) Telomerase activity assay of in vivo reconstituted human mutant telomerase. The telomerase RNA genes are expressed in vivo from phTR(wt), phTR(m2), or phTR(+2U), as indicated above the gel. A mock transfection with only phTERT-HA was included to ensure the absence of endogenous hTR expression (lanes 1,2).