Figure 8.

Fibrillarin was not recruited to the nucleolar domain when the onset of nonribosomal zygotic transcription was inhibited. The fluorescent in situ transcription assay by immunolabeling of incorporated Br-UTP coupled to immunolabeling of fibrillarin was performed on nuclei isolated 9 h after fertilization from control embryos (A) and embryos treated with actinomycin D (B and C). A, At this time, incorporation of Br-UTP was detected in the nucleoplasm as punctuated signals (a) not colocalized with fibrillarin to the nucleolar domain in control nuclei (b). This transcription signal was inhibited by incubation of nuclei in 100 μg/ml of α-amanitin before and during incorporation of Br-UTP (d). B and C, A non-RNA pol I transcription signal was also detected in nuclei isolated 9 h after fertilization when embryos were treated with 0.5 μg/ml of actinomycin D (g), but not when embryos were treated with 2 μg/ml of actinomycin D (j). In this latter case, fibrillarin was maintained in large PNBs dispersed in the nucleoplasm (k). Arrows indicate the position of the nucleolar domains in a and g. DNA staining with DAPI is shown in (c, f, i, and l). Bars, 10 μm.