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. 2009 Mar 27;284(13):8557–8566. doi: 10.1074/jbc.M806702200

FIGURE 5.

FIGURE 5.

RLIM regulates the levels of endogenous TRF1 protein. A, HT1080 clones stably expressing RLIM-V5 (OE-1 and OE-2) or the control empty vector were harvested at 75 population doublings and subjected to immunoblotting with anti-RLIM, anti-TRF1, anti-TRF2, or β-actin antibodies. The relative levels of RLIM and TRF1 were determined from three independent experiments using β-actin as a loading control. B, HT1080 clones stably expressing RLIM shRNA2 (shRNA2-1 and shRNA2-2) or scrambled shRNA were harvested at 75 population doublings and subjected to immunoblotting with anti-RLIM, anti-TRF1, anti-TRF2, or β-actin antibodies. The relative levels of RLIM and TRF1 were determined from three independent experiments using β-actin as a loading control. C, HT1080 cells stably expressing RLIM shRNA2 (shRNA2-1 and shRNA2-2) or scrambled shRNA were transfected with wild type RLIM (wt RLIM) or RNAi-R-RLIM and examined for expression of endogenous RLIM, TRF1, and TRF2 by immunoblotting. D, HT1080 cells were transiently transfected with siRNA duplexes, either nontargeting control or targeting RLIM (siRNA1 and siRNA2) for 3 days and examined for the expression of endogenous RLIM, TRF1, and TRF2 by immunoblotting. E, HT1080 cells stably expressing RLIM shRNA2 (shRNA2-1 and shRNA2-2) or scrambled shRNA were transfected with FLAG-TRF1 or FLAG-TRF2 and subjected to immunoprecipitation (IP) with anti-FLAG antibody, followed by immunoblotting with anti-RLIM antibody.