Fig. 4.

Cells with reduced Htt levels show compromised RNA-induced gene silencing. (A) The effect of Htt knockdown on silencing of a Luc reporter with Luc siRNA. HeLa cells were first transfected (Tfxn1) with control, GW182, Htt, or Ago2 siRNA, and subsequently (Tfxn2) with SV40 promoter-Luc reporter plasmid, a plasmid encoding β-galactosidase, and Luc siRNA where indicated. The ratio of luciferase activity to β-galactosidase was measured and graphed on a log scale as the percentage of expression relative to the cells transfected with the control siRNA without Luc siRNA (set to 100%) (n = 16). (B) Immunoblot analysis of cell lysates from A. (C) The effect of Htt knockdown on the ability to silence a miRNA reporter by endogenous let-7b miRNA. HeLa cells were transfected with lamin A/C, Htt, or eIF6 siRNA, and a Renilla Luc reporter that contained human let-7b element or a mutant element. Normalized ratio of Renilla Luc to SV40-firefly Luc is presented. (n = 4).