Diversity in Nuclear Pore Complexes Figure 4. Immunolabeling patterns of native NUP62 and NUP214 antibodies are consistent with those produced by V5 epitope tag antibodies detecting ectopically expressed nucleoporins. Immunofluorescent images represent 3D deconvolved projections of 250 nm optical sections through 10 to 15 um of the z axis. Projections are shown on x,y, x,z, and z,y planes, as indicated. Bar represents 10 um. A. TOV112D cells were transfected with an expression plasmid for V5 epitope-tagged NUP62 and cultured for 72 hours. The distribution of NUP62 was analyzed in pairs of post-mitotic cells expressing V5 epitope-tagged NUP62. Immunofluorescence analyses of NUP62 (green) and V5 epitope-tagged NUP62 (red) were performed with Ab1 and V5 epitope tag antibodies, respectively. Blue, DAPI stain; yellow, overlap between green and red. B. TOV112D cells were transfected with an expression plasmid for V5 epitope-tagged NUP214 and cultured for 72 hours. The distribution of NUP214 was analyzed in pairs of post-mitotic cells expressing V5 epitope-tagged NUP214. The images of the pair of post-mitotic cells shown have been cropped individually to avoid interference from surrounding cells, particularly on the x,z and z,y planes. Immunofluorescence analyses of NUP214 (red) and V5 epitope- tagged NUP214 (green) were performed with Ab4 and V5 epitope tag antibodies, respectively. Blue, DAPI stain; yellow, overlap between green and red. C. TOV112D cells were transfected with an expression plasmid for V5 epitope-tagged NUP214 and cultured for 72 hours. The distributions of NUP62 and NUP214 were analyzed in pairs of post-mitotic cells expressing V5 epitope-tagged NUP214. Immunofluorescence analyses of NUP62 (green) and V5 epitope-tagged NUP214 (red) were performed with Ab1 and V5 epitope tag antibodies, respectively. Blue, DAPI stain; yellow, overlap between green and red. doi:10.1371/journal.pone.0036137.g004 the cells in a proliferating culture (Fig. 7). Among the cells within each culture, overt changes in NPC patterning were only apparent in cells during M phase of the cell cycle. Images of the nuclei of several interphase cells were analyzed to determine radial distributions of NUP62 and NUP214 signals over multiple sectors. Integrated intensities for radial projections were normalized according to length, plotted against relative nuclear radial distance, and curves were fitted by polynomial regression. As shown in Fig. 8, differences between NUP62 and NUP214 nuclear immunolabeling patterns are readily apparent in TOV112D and PLoS ONE | www.plosone.org 6 April 2012 | Volume 7 | Issue 4 | e36137