HOXA expression signature in HOXA transformed cell lines. (A) In total 4 cell lines (2 without and 2 with coexpression of Meis1) were generated by transduction with HOXA1, HOXA4, HOXA6, and as control with HOXA9. RNA was extracted from these lines, and a complete quantitative HOXA expression profile was assembled by RT-PCR. Plotted are relative HOXA (and Meis1) expression levels for cells transduced with HOX-only (light columns) or with a HOXA/Meis1 combination (dark columns). One expression unit corresponds to a ΔC_t of 20 relative to actin. Plotted are average and SD of triplicates. (B left) Differentiation kinetics of cells transformed by conditional HOXA1 and HOXA6. A conditional derivative of HOXA1 was created by fusion with the tamoxifen responsive ligand binding domain of a modified estrogen receptor. Cell lines were generated in the presence of tamoxifen, and at time point “0” tamoxifen was withdrawn. RNA was extracted from these cells 6, 24, 48, 72, and 144 hours after tamoxifen depletion and absolute levels of endogenous (murine) Hoxa9, c-Myb, c-Kit, and Gr-1 were measured by q-RT PCR. c-Kit (CD117) is a marker for precursor cells whereas Gr-1 indicates differentiation. c-Myb has been described as an essential mediator of HOXA9 mediated transformation. Given are average values and standard deviations of triplicates of relative quantities calibrated to actin and normalized to concentrations in the presence of tamoxifen (= time point 0 hours). (B right) Similar experiment as described in the left panel, with the use of HOXA6-ER cells.