FIG. 12.

S107 plasmacytoma cells, which lack NF-κB, still exhibit prominent interactions between Igκ gene cis-acting sequences. (A) Western blot of nuclear protein extracts from MPC-11 and S107 cells showing the absence of p65NF-κB in S107 samples but not MPC-11 samples. Probing the same blot with anti-actin antibodies serves as a loading and protein integrity control. (B) Semiquantitative PCRs were performed by using 1, 3, and 9 ng of DNA templates with the indicated input, bound, and preimmune control fractions with antibodies against p65NF-κB or E47 transcription factors as indicated. (C) Comparison of S107 and MPC-11 plasmacytoma-cell-specific looping interactions between Ed and the other two enhancers. Also assayed for were interactions between the Vκ4-58 gene promoter, which is rearranged to Jκ4 in S107 cells (8, 31, 32), and Ed. Quantification of PCR signals with primer 17 as an anchor is shown. The vertical lines indicate the NspI cutting sites that were selected for analysis of the looping interactions. Data from MPC-11 is taken from Fig. 7, and the asterisks specifically indicate what interactions are statistically significant for a comparison between S107 and P815 cells from Fig. 7 data. It should be noted that NspI fragment 9 does not include the Vκ19-17 promoter from MPC-11 cells, which is on the next upstream restriction fragment (see Fig. 7).