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Fig. 7. Intranuclear distribution of RNA polymerase IIo and analysis of colocalization with HIF-1, Arnt, CBP and SRC-1. Insets show enlarged examples of the different observed foci in each image. (A) RNA polymerase IIo shows an intranuclear distribution in discrete accumulation foci. Immunocytochemistry for RNA polymerase IIo (RNA pol IIo) was performed using anti-RNA polymerase IIo antibody (CC-3) as described in Materials and Methods. (a) Enlargement of a nuclear accumulation focus of endogenous RNA polymerase IIo. (B) CBP colocalizes with RNA polymerase IIo in a large number of nuclear accumulation foci. Cells were transfected with pYFP-CBP as described in Fig. 1A, and anti-RNA polymerase IIo immunocytochemistry was performed using the CC-3 antibody. An RNA polymerase IIo accumulation focus (a), a YFP-CBP accumulation focus (b) and a focus where RNA polymerase IIo colocalized with YFP-CBP (c) are shown. (C) Colocalization of SRC-1 and RNA polymerase IIo in a few discrete foci. Cells were transfected with pYFP-SRC-1 and immunocytochemistry was performed as indicated above. Enlargement of an RNA polymerase IIo accumulation focus (a), a YFP-SRC-1 accumulation focus (b) and a focus of colocalization between RNA polymerase IIo and YFP-SRC-1 are shown. (D) The intranuclear distribution of Arnt in discrete foci does not overlap with RNA polymerase IIo. Cells were transfected with pYFP-Arnt. Enlarged images of an RNA polymerase IIo accumulation focus (a) and a YFP-Arnt accumulation focus (b) are shown. (E) The intranuclear distribution pattern of HIF-1 and Arnt partially overlaps with that of RNA polymerase IIo. HEK 293 cells were transfected with pCFP-HIF-1 (CFP-HIF-1) and pYFP-Arnt (YFP-Arnt) as indicated in the legend of Fig. 6B. An RNA polymerase IIo accumulation focus (a), a CFP-HIF-1/YFP-Arnt accumulation focus (b) and a focus of colocalization between RNA polymerase IIo and CFP-HIF-1/YFP-Arnt (c) are shown. (F) Distribution of CFP-HIF-1 and YFP-CBP into some discrete foci of colocalization with RNA polymerase IIo. Enlargements of an RNA polymerase IIo accumulation focus (a), a CFP-HIF-1/YFP-CBP accumulation focus (b) and a focus of colocalization between RNA polymerase IIo and CFP-HIF-1/YFP-CBP (c) are shown. All figures are single confocal sections obtained from HEK 293 cells treated with 100 µM CoCl₂ for 2 hours. Bar, 3 µm for all images.

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