Fig. 3. Characterization of nucleolus-associated PML structures. (A-H) Senescence-associated nuclear bodies. (A,B) Phase contrast images of WI-38 cells transduced with GFP-PML-I (see Movie 1 in supplementary material). (C-H) Untransfected WI-38 cells labelled for the following proteins: PML (green) or Sp100, DAXX, SUMO-1 and 2, Brca1 and Nbs1 (red). (I-P) Nucleolar components in SANB. (I-L) PML (green). RNA, UBF, Fibrillarin and B23 appear in red. (M,N) RNA pol I associated transcriptional activity in SANB. PML appears red and BrU green. (O,P) PML budding structures from the nucleolus. PML appears green and the nucleolus (fibrillarin) red. (Q) Biopsies from skin cancers were analysed with PML antisera. PML localises in the nucleolus (white arrow). (R,S) Senescent Idh4 cells exhibit SANB-like bodies. PML (green) and B23 or ubiquitin (red). White filled arrowhead, SANB-containing nucleolar components; open and green arrowheads, nucleolar-free SANB not in contact with nucleolus. All the images were obtained using photon microscopy, except images M-N and S, which were obtained with confocal microscopy.