FIGURE 5.

Nuclear localization of NES-deleted monomeric PCNA. A, tridimensional structure of trimeric and monomeric PCNA. Left panel: NES1 localization within the PCNA tridimensional structure (Protein Data Bank (PDB) ID: 1VYM). The interdomain connecting loop of PCNA is shown in green. The NES1 sequence is localized in the inner face of the PCNA trimer (cyan central helix). The protein is represented with graphics, and the NES1 atoms (Ile-11–Asn-23) are represented with balls colored following their type (nitrogen, blue; oxygen, red; carbon, cyan). Right panel: the Y114A mutation results in PCNA monomerization. The close-up of the mutated amino acids (orange balls) of the NES1 shows the following residues: Leu-12, Leu-16, Leu-19, and Leu-22. Both figures have been generated with PyMOL (42). B, analysis of GFP-PCNA localization by confocal scanning microscopy in HeLa cells transfected with pcDNA3-GFP-wtPCNA (wild-type PCNA), pcDNA3-GFP-PCNAY114A, pcDNA3-GFP-PCNAY114AΔ12–16NES1, or pcDNA3-GFP-PCNAΔ12–16NES1). Nuclei were visualized by Hoechst staining (original magnification ×630, scale bar = 20 μm).