Diversity in Nuclear Pore Complexes Figure 6. Total accumulation and distribution of NUP62 and NUP214 in different cell lines. Top left corner: immunoblot analyses of Hs 832(C).T (Hs), TOV112D (112D), HEK293 (293), S12, and COS7 (cos7) cell lysates. Antibodies are explained in the text. Immunofluorescent images for the indicated cell lines 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. immunofluorescence analyses of NUP62 (green) and NUP214 (red) were generated using Ab1 and Ab4, respectively. Blue, DAPI stain; yellow, overlap between green and red. Bars represent 2 um. doi:10.1371/journal.pone.0036137.g006 been reported as 125 nm [46], differing from the observed values because of the physical contributions of primary and secondary antibodies and light diffraction. Discussion Architectural diversity among NPCs could result from several mechanisms, including differential expression or distribution of nucleoporins, posttranslational modification of NPC proteins, and structural alterations associated with nucleocytoplasmic transport. Since many nucleoporins are distributed between NPCs and cytoplasmic and intranuclear cellular pools (discussed in [47]), differences in total expression may not manifest directly as differences in NPC composition. Similarly, cells with equivalent expression of a specific nucleoporin may not display equivalent distribution of that nucleoporin among NPCs, as allocation to cytoplasmic and intranuclear compartments may differ by cell type and directly impact distribution to NPCs. Posttranslation modi- fications of NUP62 and NUP214 include addition of O-linked N- acetylglucosamine [17,48,49,50] and phosphorylation [51,52,53], and may affect nucleoporin immunolabeling patterns if present on relevant epitopes in specific populations of NPCs. Alterations in NUP214 structure associated with its status in nucleocytoplasmic transport processes [32] may also affect immunolabeling. The results presented here suggest that cell-type associated differences in NUP62 and NUP214 NPC immunolabeling patterns arise from sorting of these FG-repeat nucleoporins to specific NPC populations. The co-localization of NUP62 and NUP214 in some NPCs indicates that binding of one nucleoporin antibody does not sterically preclude binding of the other to the PLoS ONE | www.plosone.org 8 April 2012 | Volume 7 | Issue 4 | e36137