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. Author manuscript; available in PMC: 2010 Mar 16.
Published in final edited form as: Cancer Cell. 2006 Nov;10(5):401–411. doi: 10.1016/j.ccr.2006.09.012

Figure 6. Trib2-dependent inhibition of C/EBPα expression and function is abrogated by siRNA-mediated knockdown of Trib2 and over-expression of C/EBPα.

Figure 6

(A–B) siRNA designed to knockdown human Trib2 (hTrib2) was electroporated into U937 cells and analyzed 24 hr later. (A) hTrib2 expression assessed by real-time RT-PCR. Error bars denote +/− S.E.M of each sample measured in triplicate. (B) Western blot of C/EBPα protein expression. Lane 1=negative control siRNA and lane 2=hTrib2 siRNA. C/EBPαp30 is undetectable in parental U937 cells.

(C–D) siRNA designed to knockdown murine Trib2 (mTrib2) was electroporated into U937 cells retrovirally expressing murine Trib2 and analyzed 24 hr later. (C) mTrib2 expression assessed by real-time RT-PCR. Error bars denote +/− S.E.M of each sample measured in triplicate. (D) Western blot of C/EBPαp42 and C/EBPαp30 protein expression. Lanes 1 & 2=U937 cells + human Trib2 siRNA and negative control siRNA respectively as in (B); lanes 3 & 4=U937-mTrib2 cells + negative control siRNA and siRNA mTrib2 respectively. ns=non-specific band

(E) 32D cells transduced with C/EBPαER (GFP)α Trib2 (tNGFR), or C/EBPαER+Trib□ were sorted for GFP and/or tNGFR expression. Sorted cells were plated in equal numbers (1x105) (day 0, 48 hrs post-transduction) in IL-3 or G-CSF +/− 1μM β-estradiol (EST). CD11b expression was assessed after 2 days. Data are representative of 3 independent experiments.