(A–F) Immuno-FISH labeling shows the active form of PolII in regions of intermingling. Serine2-phosphorylated PolII was immunolabeled with H5 antibodies and AlexaFluor488, before hybridization with paints for Chromosome 3 and all other chromosomes (WG-3). A magnified view of Chromosome 3 (A) and the remaining chromatin (B) is shown, which contains a region of intermingling (C). Active PolII is found within masks delineating Chromosome 3 territory (D), the remaining chromatin (E) and regions of intermingling (F). PolII sites are not excluded or enriched in areas of intermingling (see
Figure S4). Bar: 1 μm.
(G) Measurements of intermingling for ten chromosome pairs in control and α-amanitin–treated lymphocytes reveals changes in intermingling volumes for four of ten pairs analyzed, showing that the extent of intermingling is affected by ongoing transcription.
(H) Differences in intermingling are not due to altered CT volumes, as these did not change after treatment with α-amanitin for 12 of 13 chromosomes analyzed. Error bars represent standard deviations (*
p < 0.05, **
p < 0.01).