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. 2006 Feb;26(3):810–821. doi: 10.1128/MCB.26.3.810-821.2006

FIG. 2.

FIG. 2.

Inducible human E47 rescues Igκ germ line transcription and recombination. (A) E47R is sufficient to activate Igκ germ line transcription. Total RNA was isolated from E2A+/+ or E2A−/− Ab-MuLV pre-B cells treated with 1 μM tamoxifen (tamox) and E47R-reconstituted Ab-MuLV B cells treated with 0.1% DMSO or 1 μM tamoxifen for 6 h. Semiquantitative RT-PCR was performed to determine the relative level of Igκ germ line transcription (Igκ-GL). EF1α serves as a positive loading control, and results are presented as threefold serial dilutions. (B) E47R promotes Igκ recombination. E47R-reconstituted pre-B cells were treated with 1 μM tamoxifen or 0.1% DMSO for 8 or 24 h. Genomic DNA was prepared from the cells, and the accumulation of Jκ1 signal break ends (SBE) was determined by LM-PCR. PCR amplification of the CD14 gene serves as a control for DNA loading and quality. (C) Igκ germ line transcription is dependent on E2A. E47R-reconstituted pre-B cells were treated with 1 μM tamoxifen for 6 h, washed, and cultured without tamoxifen. Total RNA was prepared at the indicated times after the removal of tamoxifen. Mock-treated cells were cultured in the presence of 0.1% DMSO for the entire duration of the experiment. Relative abundance of Igκ germ line transcript and EF1α transcript was determined by RT-PCR.