Figure 3.
The TRD of MeCP2 represses L1 retrotransposition. (A) Reporter construct JM101/L1.2ΔCMV or G5JM101/L1.2ΔCMV was co-transfected with the empty vector pcDNA1.1 or with expression construct Gal4-TRDMeCP2 into HeLa cells. Selection with G418 began 3 days after transfection and, after 12 days, G418 resistant foci were fixed and stained with Giemsa for visualization. Results of a representative transposition assay are shown. (B) Effect of Gal4-TRDMeCP2 on relative transposition frequencies of the neo-marked L1.2 reporters. The bar chart represents the data in Table 1A. Relative retrotransposition frequencies refer to the mean retrotransposition rate of construct JM101/L1.2ΔCMV (357 ± 17 × 106), which is set as 1.0. (C) Expression of Gal4-TRDMeCP2 72 h post-transfection was controlled for by immunoblot analysis with anti-Gal4BD antibody. The upper 57-kDa band represents the full-length Gal4-TRDMeCP2. The lower band very likely results from cellular protease activity, since the C-terminal half of MeCP2 exhibits prominent sensitivity to proteolysis (10).