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. 2010 Jun 8;38(19):6673–6683. doi: 10.1093/nar/gkq501

Figure 3.

Figure 3.

Mutation of the conserved amino acid residues abolish repression by NED. (A) Schematic representation of the NED region and its alignment across the species (Dm: Drosophila melanogaster, Da: Drosophila ananassae; Ap: Apis mellifera; TNRC6A, B, and C are human GW182 homologs). Alignment was performed with a T-Coffee tool [(41) see Supplementary Figure S1 for full alignment]. Mutated residues (always to alanine) are shown in red. The numbers correspond to amino acid position. Asterisks mark residues identical in all sequences, colons mark conservative substitutions, dots mark semi-conservative substitutions. (B) Repression of FLuc-boxB mRNA by the NED and its point mutants. Drosophila S2 cells were co-transfected with plasmids encoding FLuc-boxB, RLuc, and either NHA-205-490 (NED) or one of its point mutants depicted in (A). For other details, see legend to Figure 1. (C) Expression levels of HA-fusion proteins as estimated by western blotting with the anti-HA antibody.