FIGURE 3.

THP1–155 cells overexpressing miR-155 showed down-regulation of PU.1 protein expression. A, THP1–155 cells were treated or not with doxycycline (DOX) to de-repress or not the expression of the miR-155 transgene, respectively. Cells were collected at different time points, and subjected to RNA extraction. RNA from these samples was subjected to specific microRNA RT qPCR, and miR-155 levels were quantified. RNU6B was employed for normalization purposes. B, protein extracts from the same samples were subjected to Western blotting, and PU.1 levels were determined. As a control, α-tubulin was also detected in the same blot. Values represent percentage of PU.1 normalized against α-tubulin and compared with control. C, the same RNA extracts were also used to perform standard RT qPCR, to detect PU.1 mRNA levels. Values were normalized against ACT1B. This figure shows one representative experiment of three. Data in A and C represent mean ± S.D. (error bars). Differences in B (+Dox versus −Dox), not significant.