FIGURE 3.

Disrupting PinX1 binding to TRF1 by the L291E mutation abolishes the ability of PinX1 to localize to telomeres in human cells. A, the L291E mutation completely abolishes the ability of either full-lengthPinX1 or its TID to interact with TRF1. The L291E mutation was introduced into full-length PinX1 or its TID, and mutant proteins were expressed in bacteria and purified as GST fusion proteins followed by GST pulldown with in vitro synthesized ³⁵S-TRF1. B–E, the L291E mutation completely abolishes the ability of either full-length PinX1 or its TID to localize to telomeres. HT1080 cells were co-transfected overnight with vectors expressing RFP-tagged TRF1 and GFP-tagged wild-type full-length PinX1 (B) or its TID (D) or their L291E point mutants (C and E) followed by subjecting them to fluorescence microscopy. Yellow arrows point to colocalization at telomeres, whereas red arrows point to colocalization at nucleoli.