Figure 6.

Transcriptional activity in LB1 silenced cells. (A) Total fluorescence intensities of Pol II, Pol IIo S2, and Pol IIo S5 in nuclei of control and LB1 silenced cells were measured using wide field microscopy. In each case, the fluorescence intensity in 40–150 nuclei was determined in each of two experiments. The relative fluorescence intensities in control cells and LB1 silenced cells with normal meshes (<1.3 μm²) and large meshes (>1.3 μm²) are plotted in the histogram as green, yellow, and red bars, respectively. Error bars represent ±SD. The bars with an asterisk mark experiments with significant changes (P < 0.05) in relative fluorescence intensity of Pol II and Pol IIo in LB1 silenced cells compared with those of controls. (B–E) Total Pol II in a nucleus with one bleb. Pol II is present both in the bleb and throughout the main nuclear body. (F–I) Pol II hyperphosphorylated on Ser 2 in the CTD (Pol IIo S2), LB1, and DNA showing the enrichment of Pol IIo in the bleb. (J–Q) RNA synthesis was visualized by BrU incorporation. Note the virtual absence of BrU incorporation into the bleb region (J–M), and the incorporation of BrU into a nucleolus in one of the few nuclei in which a nucleolus has entered the bleb (N–Q). (R–U) Localization of H3K4, LB1, and DNA demonstrating that this histone mark is present both in blebs and the nuclear body. (V–Y) Localization of H3K36, LB1, and DNA showing the absence of this histone mark within blebs. Bars, 5 μm.