Nucleolar segregation is observed upon rDNA transcriptional arrest either in physiological conditions or induced Fig. 2 The subnucleolar constituents revealed by Xuorescence microscopy. (A) The rDNA transcription machinery, illustrated by UBF labeling, is localized in several foci corresponding to FCs in HeLa cells (a). rDNA transcription sites detected by in situ BrUTP incorporation (b), mainly colocalize with UBF as seen by the merge (c). The nucleus is visualized by Dapi staining (d). (B) HeLa cells expressing either Wbrillarin-GFP fusion or DsRed-B23 fusion. Fibrillarin decorates the DFC (a, b) whereas B23 decorates the GC (c, d). In ActDtreated cells nucleoli are segregated, Wbrillarin localizes in caps (e, f) contrary to B23 that localizes in the central body and outside the caps (g, h). Arrowheads point the caps. Bars: 10 m by low doses of actinomycin D (ActD). The segregation of nucleoli is characterized by the separation of the nucleolar components that remain close to each other but no longer intermingle ( Fig. 2Be -h) (for reviews see Hadjiolov 1985; Hernandez-Verdun and Junéra 1995; Scheer and Benavente 1990) . The eVect of ActD on nucleolar organization follows sequential changes: Wrst the Wbrillar components (FCs and DFC) condense and migrate towards the periphery of the nucleolus, after which the nucleolar components segregate to Wnally form a central body associated with caps (Hadjiolova et al. 1995) . In the caps are several proteins related to the RNA polymerase (pol) I transcription machinery such as UBF, close to Wbrillarin-containing caps. In the central body are proteins derived from the GC, some of which are progressively released, such as PM-Scl 100. It was recently demonstrated that certain nucleolar caps of segregated nucleoli could recruit factors involved in mRNA splicing. In this case, localization is induced by inhibition of both RNA pol I (rRNA transcription arrest) and RNA pol II (mRNA transcription arrest) (Shav-Tal et al. 2005 ). This is not observed when only RNA pol I is inhibited indicating that the composition of a segregated nucleolus can be more complex when induced by general transcription inhibition.