TIA1 and G3BP1 are required for SAMHD1 to inhibit LINE-1 retrotransposition.
(A) Knockdown of TIA1 or G3BP1 prevents the SAMHD1 from inhibiting LINE-1. HeLa cells were transfected with siRNA targeting TIA1 or G3BP1 prior to co-transfection with Myc-SAMHD1 and CMV-L1-neoRT. Neomycin-resistant colonies were scored and results of three independent experiments are shown in the bar graph. Levels of TIA1, G3BP1 and Myc-SAMHD1 were determined by western blotting. The knockdown efficiency of TIA1 and G3BP1 was calculated on the basis of the intensities of TIA1 or G3BP1 bands in the western blots. (B) Endogenous SAMHD1 loses inhibition of LINE-1 upon depletion of TIA1 or G3BP1. HeLa cells were treated with siRNAs targeting TIA1, G3BP1 or SAMHD1, followed by transfection with CMV-L1-neoRT DNA. Number of neomycin-resistant colonies was determined and shown in the bar graph. The number of neomycin-resistant colonies with control siRNA is arbitrarily set as 1. Levels of endogenous TIA1, G3BP1 and SAMHD1 were examined by western blotting. The knockdown efficiency was calculated on the basis of the protein band intensities as determined using the ImageJ program. * indicates p< 0.05, ns denotes “not significant”.