Nuclear Distributions of NUP62 and NUP214 Suggest Architectural Diversity and Spatial Patterning among Nuclear Pore Complexes Yayoi Kinoshita1, Tamara Kalir1, Peter Dottino 3 , D. Stave Kohtz1,2* 1 Department of Pathology, Mount Sinai School of Medicine, New York, New York, United States of America, 2 Department of Oncological Sciences, Mount Sinai School of Medicine, New York, New York, United States of America, 3 Obstetrics, Gynecology, and Reproductive Science, Mount Sinai School of Medicine, New York, New York, United States of America Abstract The shape of nuclei in many adherent cultured cells approximates an oblate ellipsoid, with contralateral flattened surfaces facing the culture plate or the medium. Observations of cultured cell nuclei from orthogonal perspectives revealed that nucleoporin p62 (NUP62) and nucleoporin 214 (NUP214) are differentially distributed between nuclear pore complexes on the flattened surfaces and peripheral rim of the nucleus. High resolution stimulated emission depletion (STED) immunofluorescence microscopy resolved individual NPCs, and suggested both heterogeneity and microheterogeneity in NUP62 and NUP214 immunolabeling among in NPC populations. Similar to nuclear domains and interphase chromosome territories, architectural diversity and spatial patterning of NPCs may be an intrinsic property of the nucleus that is linked to the functions and organization of underlying chromatin. Citation: Kinoshita Y, Kalir T, Dottino P, Kohtz DS (2012) Nuclear Distributions of NUP62 and NUP214 Suggest Architectural Diversity and Spatial Patterning among Nuclear Pore Complexes. PLoS ONE 7(4): e36137. doi:10.1371/journal.pone.0036137 Editor: Michael Klymkowsky, University of Colorado, Boulder, United States of America Received March 3, 2011; Accepted April 2, 2012; Published April 27, 2012 Copyright: 2012 Kinoshita et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was funded by a generous grant from the Ruttenberg family, and by National Institutes of Health/National Cancer Institute (NIH/NCI) grant number 1R03CA141318 (to DSK). Widefield, confocal laser scanning, and STED microscopy were performed at the MSSM-Microscopy Shared Resource Facility, which is supported by NIH/NCI shared resources grant 5R24 CA095823-04, NSF Major Research Instrumentation grant DBI-9724504, and NIH shared instrumentation grant 1 S10 RR0 9145-01. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: stave.kohtz@mssm.edu Introduction Nuclear pore complexes (NPC) assemble in the nuclear envelope (NE) from more than 30 different proteins, and are organized into multiprotein subcomplexes. Multiples of eight units of each subcomplex assemble into the modular structures of the pore, which include a symmetric pair of core inner ring complexes, an asymmetric pair of annular rings on the cytoplasmic and nuclear faces, and asymmetric filamentous structures projecting into the nucleoplasm and cytoplasm [1,2]. In vertebrates, the conserved NUP107-160 subcomplex forms the essential symmetric pair of inner ring complexes of the pore (reviewed in [3,4]). During assembly of nascent pores, the NUP107-160 subcomplex is recruited to the nuclear envelope by ELYS/MEL28, a chromatin binding protein [5], and the ELYS-NUP107-160-chromatin complex recruits membrane vesicles containing transmembrane proteins that will anchor the NPC into the nuclear envelope [6]. Each of three transmembrane glycoproteins (POM121, gp210, and NDC1) or combinations thereof can anchor the pore complex in the NE, and their expression varies between cell types and species [7,8]. Located symmetrically on both the cytoplasmic and nuclear faces of the NPC, NUP155 interacts with Gle1 and nucleoporin pCG1 to mediate mRNA transport [9,10]. Residing near the nuclear envelope and interacting with lamin B, NUP53 associates with a putative NPC complex containing NUP93, NUP155 and NUP205 [11]. Proteolytic cleavage of a 186 kDa precursor protein results in the generation of NUP98, an FG-repeat containing nucleoporin [12], which resides on both the nuclear and cytoplasmic sides of the NPC [13], and of NUP96, for which evidence has been presented of a role in core inner ring assemble [14]. The NUP62 subcomplex, which contains the FG-repeat nucleoporin NUP62 [15], forms rings on the inner channel of the pore [13,16]. The cytoplasmic or nuclear annular rings contain eight subunits of NUP88-NUP214 [17,18,19] or NUP153 [20], respectively. Filamentous structures projecting into the cytoplasm or nucleus are formed by NUP153/Tpr [20] or NUP358 [21], respectively (reviewed in [4]). Export of mRNA from the nucleus is a complex process that in yeast and probably in metazoans is linked to mRNA processing. Export proteins that interact with the NPC and mediate transport recognize adaptors that bind maturing RNA, and many of these adaptors are directly involved in RNA processing (reviewed in [22,23]). Passage of protein cargo through the NPC is mediated by carrier proteins (karyopherins), which include the importins, exportins, and transportins, and differential expression of the karyopherins is an important determinant in commitment to specific cell lineages (reviewed in [24]). Cargo/karyopherin complexes transported from the cytoplasm to the nucleus dissociate in the nucleus by interacting with RanGTP (reviewed in [25]). Likewise, karyopherin- or cargo/karyopherin-RanGTP complexes pass to the cytoplasm, where RanGAP promotes PLoS ONE | www.plosone.org 1 April 2012 | Volume 7 | Issue 4 | e36137