Let-7a is down-regulated during transformation by NF-κB.
(A) RNA levels of individual let-7 family members in cells treated with TAM for the indicated times.
(B) Levels of let-7a RNA in untreated and TAM-treated (36h) cells in the presence or absence of NF-κB inhibitor (5 μM BAY-117082 and 6 μM JSH-23).
(C) Lin28B protein levels (western blot) in ER-Src cells at the indicated times after TAM treatment.
(D) Lin28B mRNA levels (mean ± SD) during transformation in the presence or absence of an NF-κB inhibitor (5 μM BAY-117082).
(E) Levels of precursor RNAs (pri-let-7a and pri-let-7d/f) in untreated and TAM-treated cells.
(F) NF-κB occupancy (fold-enrichment) at the Lin-28B and let-7a loci as determined by chromatin immunoprecipitation of crosslinked cells that were or were not treated with TAM.
(G) Luciferase activity (mean ± SD)) of Lin28B vector containing the NF-κB binding site during transformation.
(H) Luciferase activity of Lin28B vector (wt, mutated or deleted in NF-κB site) in TAM-treated (36h) cells.
(I) Anchorage-independent growth assays (microscopic counting of 50 μm colonies) of untreated and TAM-treated cells transfected with the indicated let-7 family members or siRNA against Lin28B.