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Super-resolution microscopy, and specifically single-molecule localization microscopy (SMLM), is becoming a transformative technology for cell biology, as it allows the study of cellular structures with nanometer resolution. Here, we review a wide range of data analyses approaches for SMLM that extract quantitative information about the distribution, size, shape, spatial organization, and stoichiometry of macromolecular complexes to guide biological interpretation. We present a case study using the nuclear pore complex as an example that highlights the power of combining complementary approaches by identifying its symmetry, ringlike structure, and protein copy number. In face of recent technical and computational advances, this review serves as a guideline for selecting appropriate analysis tools and controls to exploit the potential of SMLM for a wide range of biological questions. Technical advances in SMLM provide nanoscale details of subcellular structures in 3D, multicolor, and with increased throughput. Analysis methods specific for SMLM extract quantitative information from the primary coordinate-based data, including underlying geometry, topology, and number of proteins. Multicolor SMLM analysis approaches reveal the relative organization and molecular interactions of proteins in subcellular structures. [ABSTRACT FROM AUTHOR]

Tau is a cytosolic microtubule binding protein that is highly abundant in the axons of the central nervous system. However, alternative functions of tau also in other cellular compartments are suggested, for example, in the nucleus, where interactions of tau with specific nuclear entities such as DNA, the nucleolus, and the nuclear envelope have been reported. We would like to review the current knowledge about tau–nucleus interactions and lay out possible neurotoxic mechanisms that are based on the (pathological) interactions of tau with the nucleus. [ABSTRACT FROM AUTHOR]

The nuclear envelope delineates the eukaryotic cell nucleus. The membrane system of the nuclear envelope consists of an outer nuclear membrane and an inner nuclear membrane separated by a perinuclear space. It serves as more than just a static barrier, since it regulates the communication between the nucleoplasm and the cytoplasm and provides the anchoring points where chromatin is attached. Fewer nuclear envelope proteins have been identified in plants in comparison with animals and yeasts. Here, we review the current state of knowledge of the nuclear envelope in plants, focusing on its role as a chromatin organizer and regulator of gene expression, as well as on the modifications that it undergoes to be efficiently disassembled and reassembled with each cell division. Advances in knowledge concerning the mitotic role of some nuclear envelope constituents are also presented. In addition, we summarize recent progress on the contribution of the nuclear envelope elements to telomere tethering and chromosome dynamics during the meiotic division in different plant species. [ABSTRACT FROM AUTHOR]

In the ongoing process of uncovering molecular abnormalities in neurodegenerative diseases characterized by toxic protein aggregates, nucleo-cytoplasmic transport defects have an emerging role. Several pieces of evidence suggest a link between neuronal protein inclusions and nuclear pore complex (NPC) damage. These processes lead to oxidative stress, inefficient transcription, and aberrant DNA/RNA maintenance. The clinical and neuropathological spectrum of NPC defects is broad, ranging from physiological aging to a suite of neurodegenerative diseases. A better understanding of the shared pathways among these conditions may represent a significant step toward dissecting their underlying molecular mechanisms, opening the way to a real possibility of identifying common therapeutic targets. [ABSTRACT FROM AUTHOR]

Nuclear pore complexes (NPCs) are large multi-protein complexes that control bidirectional trafficking of macromolecules between the nucleus and cytoplasm. This trafficking is highly regulated and participates in a considerably broader range of cellular activities, including defense responses against pathogens in plants. Recently, NPC is emerging as a platform to physically associate the underlying chromatin with the nuclear periphery, thus regulating chromatin structure and gene expression. For instance, NPC components have been shown to promote the formation of specific genomics loops, which is linked to transcriptional memory for rapid reactivation of genes. With newly developed techniques and tools, our insight in this area has been substantially advanced. This review summarizes recent works on the molecular function of NPC machinery as hubs for transcriptional regulation and compares systems between plant and non-plant organisms. [ABSTRACT FROM AUTHOR]

Background: Starch is synthesized during daylight for temporary storage in leaves and then degraded during the subsequent night to support plant growth and development. Impairment of starch degradation leads to stunted growth, even senescence and death. The nuclear pore complex is involved in many cellular processes, but its relationship with starch degradation has been unclear until now. We previously identified that two Nucleoporin98 genes (Nup98a and Nup98b) redundantly regulate flowering via the CONSTANS (CO)-independent pathway in Arabidopsis thaliana. The double mutant also shows severe senescence phenotypes. Results: We find that Nucleoporin 98 participates in the regulation of sugar metabolism in leaves and is also involved in senescence regulation in Arabidopsis. We show that Nup98a and Nup98b function redundantly at different stages of starch degradation. The nup98a-1 nup98b-1 double mutant accumulates more starch, showing a severe early senescence phenotype compared to wild type plants. The expression of marker genes related to starch degradation is impaired in the nup98a-1 nup98b-1 double mutant, and marker genes of carbon starvation and senescence express their products earlier and in higher abundance than in wild type plants, suggesting that abnormalities in energy metabolism are the main cause of senescence in the double mutant. Addition of sucrose to the growth medium rescues early senescence phenotypes of the nup98a-1 nup98b-1 mutant. Conclusions: Our results provide evidence for a novel role of the nuclear pore complex in energy metabolism related to growth and development, in which Nup98 functions in starch degradation to control growth regulation in Arabidopsis. [ABSTRACT FROM AUTHOR]

Nuclear pore complexes (NPCs) are main channels controlling nucleocytoplasmic transport and are composed of approximately 30 nucleoporins (NUPs). Emerging evidence suggests that some NUP genes have specialized functions that challenge the traditional view of NPCs as structures of uniform composition. Here, we analysed the role of six outer‐ring components of NPC at normal and warm growth temperatures by examining their loss‐of‐function mutants in Arabidopsis thaliana. All six NUP subunits, NUP85, NUP96, NUP 133, NUP 160, SEH1 and HOS1, have a non‐redundant temperature‐influenced function in one or more of the processes, including rosette growth, leaf architecture and intracellular immune receptor‐mediated disease resistance. At the molecular level, NUP85 and NUP133 are required for mRNA export only at warm temperature and play a larger role in the localization of transcription factor at warm temperature. In addition, NUP96 and HOS1 are essential for the expression of high temperature‐responsive genes, which is correlated with their larger activity in facilitating nuclear accumulation of the transcription factor PIF4 at warm temperature. Our results show that subunits of NPC have differential roles at different temperatures, suggesting the existence of temperature‐influenced NPC complexes and activities. Functions of six outer‐ring subunits of a nuclear pore complex are examined in Arabidopsis thaliana under normal and elevated temperatures. The temperature‐influenced roles of these subunits suggest the existence of temperature‐influenced nuclear pore complexes and activities. [ABSTRACT FROM AUTHOR]

The nuclear periphery is a hotspot for the accumulation of age-induced damage in eukaryotic cells. The types of damage that occur at the periphery and their phenotypic consequences have begun to be characterized; however, the mechanisms by which cells repair or eliminate nuclear damage remain poorly understood. Using budding yeast meiosis as a natural system to study cellular rejuvenation, we recently discovered a novel nuclear quality control event, in which age-induced damage is sequestered away from dividing chromosomes to a discarded nuclear compartment that we term the GUNC (for "Gametogenesis Uninherited Nuclear Compartment"). Interestingly, extensive nuclear remodeling occurs even in young cells, including a surprising modularity of the nuclear pore complex, suggesting a general contribution to gamete fitness. In this review, we discuss these findings in the context of recent evidence that the nuclear periphery is a highly dynamic region critical for cellular health. [ABSTRACT FROM AUTHOR]

Several aggregation-prone RNA-binding proteins, including FUS, EWS, TAF15, hnRNP A1, hnRNP A2, and TDP-43, are mutated in neurodegenerative diseases. The nuclear–cytoplasmic distribution of these proteins is controlled by proteins in the karyopherin family of nuclear transport factors (Kaps). Recent studies have shown that Kaps not only transport these proteins but also inhibit their self-association/aggregation, acting as molecular chaperones. This chaperone activity is impaired for disease-causing mutants of the RNA-binding proteins. Here, we review physical data on the mechanisms of self-association of several disease-associated RNA-binding proteins, through liquid–liquid phase separation and amyloid fiber formation. In each case, we relate these data to biophysical, biochemical, and cell biological data on the inhibition of self-association by Kaps. Our analyses suggest that Kaps may be effective chaperones because they contain large surfaces with diverse physical properties that enable them to engage multiple different regions of their cargo proteins, blocking self-association. [ABSTRACT FROM AUTHOR]

Nucleoporins (NUPs) are an essential component of the nuclear-pore complex, which regulates nucleocytoplasmic transport of macromolecules. Pathogenic variants in NUP genes have been linked to several inherited human diseases, including a number with progressive neurological degeneration. We present six affected individuals with bi-allelic truncating variants in NUP188 and strikingly similar phenotypes and clinical courses, representing a recognizable genetic syndrome; the individuals are from four unrelated families. Key clinical features include congenital cataracts, hypotonia, prenatal-onset ventriculomegaly, white-matter abnormalities, hypoplastic corpus callosum, congenital heart defects, and central hypoventilation. Characteristic dysmorphic features include small palpebral fissures, a wide nasal bridge and nose, micrognathia, and digital anomalies. All affected individuals died as a result of respiratory failure, and five of them died within the first year of life. Nuclear import of proteins was decreased in affected individuals' fibroblasts, supporting a possible disease mechanism. CRISPR-mediated knockout of NUP188 in Drosophila revealed motor deficits and seizure susceptibility, partially recapitulating the neurological phenotype seen in affected individuals. Removal of NUP188 also resulted in aberrant dendrite tiling, suggesting a potential role of NUP188 in dendritic development. Two of the NUP188 pathogenic variants are enriched in the Ashkenazi Jewish population in gnomAD, a finding we confirmed with a separate targeted population screen of an international sampling of 3,225 healthy Ashkenazi Jewish individuals. Taken together, our results implicate bi-allelic loss-of-function NUP188 variants in a recessive syndrome characterized by a distinct neurologic, ophthalmologic, and facial phenotype. [ABSTRACT FROM AUTHOR]

The nuclear envelope (NE) is the central organizing unit of the eukaryotic cell serving as a genome protective barrier and mechanotransduction interface between the cytoplasm and the nucleus. The NE is mainly composed of a nuclear lamina and a double membrane connected at specific points where the nuclear pore complexes (NPCs) form. Physiological aging might be generically defined as a functional decline across lifespan observed from the cellular to organismal level. Therefore, during aging and premature aging, several cellular alterations occur, including nuclear‐specific changes, particularly, altered nuclear transport, increased genomic instability induced by DNA damage, and telomere attrition. Here, we highlight and discuss proteins associated with nuclear transport dysfunction induced by aging, particularly nucleoporins, nuclear transport factors, and lamins. Moreover, changes in the structure of chromatin and consequent heterochromatin rearrangement upon aging are discussed. These alterations correlate with NE dysfunction, particularly lamins' alterations. Finally, telomere attrition is addressed and correlated with altered levels of nuclear lamins and nuclear lamina‐associated proteins. Overall, the identification of molecular mechanisms underlying NE dysfunction, including upstream and downstream events, which have yet to be unraveled, will be determinant not only to our understanding of several pathologies, but as here discussed, in the aging process. [ABSTRACT FROM AUTHOR]

Cardiovascular disease is a pressing health problem with significant global health, societal, and financial burdens. Understanding the molecular basis of polygenic cardiac pathology is thus essential to devising novel approaches for management and treatment. Recent identification of uncharacterized regulatory functions for a class of nuclear envelope proteins called nucleoporins offers the opportunity to understand novel putative mechanisms of cardiac disease development and progression. Consistent reports of nucleoporin deregulation associated with ischemic and dilated cardiomyopathies, arrhythmias and valvular disorders suggests that nucleoporin impairment may be a significant but understudied variable in cardiopathologic disorders. This review discusses and converges existing literature regarding nuclear pore complex proteins and their association with cardiac pathologies, and proposes a role for nucleoporins as facilitators of cardiac disease. • The nuclear pore complex regulates development and disease of multiple systems via a variety of mechanisms. • Identification and consistent demonstration of nucleoporin dysfunction associated with clinical cardiac diseases. • Nucleoporins have potential roles in cardiomyopathy, arrhythmogensis, and congenital heart disease. • Molecular profiling facilitates systems biology understanding of relationships between nucleoporins and cardiopathogenesis. [ABSTRACT FROM AUTHOR]

Influenza A viruses have a single-stranded RNA genome consisting of 8 segments. Each RNA segment associates with the nucleoprotein (NP) and viral RNA polymerase to and from a viral ribonucleoprotein (vRNP) particle. The viral mRNA synthesis is dependent on a capped primer derived from nascent host RNA transcripts. For these processes to take place, vRNPs must pass through the cell nuclear pore complex (NPC) to the nucleus. The influenza A virus NS2 protein, also called the nuclear export protein (NES), has an important role in the nucleocytoplasmic transport of vRNPs. This protein interacts with the host cellular nucleoporins during the nuclear export of vRNPs. In this study, the human nucleoporin 214 (Nup214) was identified as an NS2-binding protein by using a yeast two-hybrid assay. The interaction between NS2 and human Nup214 was confirmed in both yeast and mammalian cells. It has been shown that the NS2 protein interacts with the amino terminal FG domain of the Nup214 protein. The influenza viral replication was suppressed in knockdown cells for the Nup214 protein. It was concluded that the FG domains of nucleoporins have an important role in the interaction of the influenza NS2 protein with host NPC for vRNA export. [ABSTRACT FROM AUTHOR]

In addition to their role in regulating transport across the nuclear envelope, increasing evidence suggests nuclear pore complexes (NPCs) function in regulating gene expression. For example, the induction of certain genes (e.g., yeast INO1) is accompanied by their movement from the nuclear interior to NPCs. As sumoylation has been linked to the regulation of chromatin spatial organization and transcriptional activity, we investigated the role of sumoylation in the expression and NPC recruitment of the INO1 gene. We observed that induction of INO1 is accompanied by both increased and decreased sumoylation of proteins associated with specific regions along the INO1 locus. Furthermore, we show that the E3 ligase Siz2/Nfi1 is required for targeting the INO1 locus to the NPC where it interacts with the SUMO isopeptidase Ulp1. Our data suggest that this interaction is required for both the association of INO1 with the NPC and for its normal expression. These results imply that sumoylation is a key regulator of INO1 targeting to the NPC, and a cycle of sumoylation and NPC-associated desumoylation events contribute to the regulation of INO1 expression. [ABSTRACT FROM AUTHOR]

Neuronal development and memory consolidation are conserved processes that rely on nuclear-cytoplasmic transport of signaling molecules to regulate gene activity and initiate cascades of downstream cellular events. Surprisingly, few reports address and validate this widely accepted perspective. Here we show that Importin-α2 (Imp-α2), a soluble nuclear transporter that shuttles cargoes between the cytoplasm and nucleus, is vital for brain development, learning and persistent memory in Drosophila melanogaster. Mutations in importin-α2 (imp-α2, known as Pendulin or Pen and homologous with human KPNA2) are alleles of mushroom body miniature B (mbmB), a gene known to regulate aspects of brain development and influence adult behavior in flies. Mushroom bodies (MBs), paired associative centers in the brain, are smaller than normal due to defective proliferation of specific intrinsic Kenyon cell (KC) neurons in mbmB mutants. Extant KCs projecting to the MB β-lobe terminate abnormally on the contralateral side of the brain. mbmB adults have impaired olfactory learning but normal memory decay in most respects, except that protein synthesis-dependent long-term memory (LTM) is abolished. This observation supports an alternative mechanism of persistent memory in which mutually exclusive protein-synthesis-dependent and -independent forms rely on opposing cellular mechanisms or circuits. We propose a testable model of Imp-α2 and nuclear transport roles in brain development and conditioned behavior. Based on our molecular characterization, we suggest that mbmB is hereafter referred to as imp-α2mbmB. [ABSTRACT FROM AUTHOR]

Defects in nucleocytoplasmic transport have been associated with several neurodegenerative disorders and, in particular, the formation of pathological protein aggregates characteristic for the respective disease. However, whether impaired nucleocytoplasmic transport is a consequence of such aggregates or rather contributes to their formation is still mostly unclear. In this review, we summarize recent findings how both soluble and stationary components of the nucleocytoplasmic transport machinery are altered in neurodegenerative diseases, in particular amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), Alzheimer's disease (AD) and Huntington's disease (HD). We discuss the functional significance of the observed defects for nucleocytoplasmic transport of proteins and mRNAs. Moreover, we highlight interesting parallels observed in physiological ageing and the premature ageing syndrome progeria and propose that they that might provide mechanistic insights also for neurodegenerative processes. [ABSTRACT FROM AUTHOR]

The spatial organization of the genome contributes to essential functions such as transcription and chromosome integrity maintenance. The principles governing nuclear compartmentalization have been the focus of considerable research over the last decade. In these studies, the genome–nuclear structure interactions emerged as a main driver of this particular 3D genome organization. In this review, we describe the interactions between the genome and four major landmarks of the nucleus: the nuclear lamina, the nuclear pores, the pericentromeric heterochromatin and the nucleolus. We present the recent studies that identify sequences bound to these different locations and address the tethering mechanisms. We give an overview of the relevance of this organization in development and disease. Finally, we discuss the dynamic aspects and self-organizing properties that allow this complex architecture to be inherited. [ABSTRACT FROM AUTHOR]

Organelle-specific nanocarriers (NCs) are highly sought after for delivering therapeutic agents into the cell nucleus. This necessitates nucleocytoplasmic transport (NCT) to bypass nuclear pore complexes (NPCs). However, little is known as to how comparably large NCs infiltrate this vital intracellular barrier to enter the nuclear interior. Here, we developed nuclear localization signal (NLS)-conjugated polymersome nanocarriers (NLS-NCs) and studied the NCT mechanism underlying their selective nuclear uptake. Detailed chemical, biophysical, and cellular analyses show that karyopherin receptors are required to authenticate, bind, and escort NLS-NCs through NPCs while Ran guanosine triphosphate (RanGTP) promotes their release from NPCs into the nuclear interior. Ultrastructural analysis by regressive staining transmission electron microscopy further resolves the NLS-NCs on transit in NPCs and inside the nucleus. By elucidating their ability to utilize NCT, these findings demonstrate the efficacy of polymersomes to deliver encapsulated payloads directly into cell nuclei. [ABSTRACT FROM AUTHOR]

Ran is a small GTPase whose nucleotide-bound forms cycle through nuclear pore complexes (NPCs) to direct nucleocytoplasmic transport (NCT). Generally, Ran guanosine triphosphate (RanGTP) binds cargocarrying karyopherin receptors (Kaps) in the nucleus and releases them into the cytoplasm following hydrolysis to Ran guanosine diphosphate (RanGDP). This generates a remarkably steep Ran gradient across the nuclear envelope that sustains compartment-specific cargo delivery and accumulation. However, because NPCs are permeable to small molecules of comparable size, it is unclear how an uncontrolled mixing of RanGTP and RanGDP is prevented. Here, we find that an NPCenriched pool of karyopherin subunit beta 1 (KPNB1, hereafter referred to as Kapß1) selectively mediates Ran diffusion across the pore but not passive molecules of similar size (e.g. GFP). This is due to RanGTP having a stronger binding interaction with Kapß1 than RanGDP. For this reason, the RanGDP importer, nuclear transport factor 2, facilitates the return of RanGDP into the nucleus following GTP hydrolysis. Accordingly, the enrichment of Kapß1 at NPCs may function as a retention mechanism that preserves the sharp transition of RanGTP and RanGDP in the nucleus and cytoplasm, respectively. [ABSTRACT FROM AUTHOR]

Nuclear pore complexes (NPCs) are protein assemblies that form channels across the nuclear envelope to mediate communication between the nucleus and the cytoplasm. Additionally, NPCs interact with chromatin and influence the position and expression of multiple genes. Interestingly, the composition of NPCs can vary in different cell-types, tissues, and developmental states. Here, we review recent findings suggesting that modifications of NPC composition, including post-translational modifications, play an instructive role in cell fate establishment. In particular, we focus on the role of cell-specific NPC deacetylation in asymmetrically dividing budding yeast, which modulates transport-dependent and transport-independent NPC functions to determine the time of commitment to a new division cycle in daughter cells. By modulating protein localization and gene expression, NPCs are therefore emerging as central regulators of cell identity. [ABSTRACT FROM AUTHOR]

In this study, we examined how channel‐forming subunits of the nuclear pore complex (NPC) are assembled into a selective channel within a highly structured scaffold ring during postmitotic assembly. We focused on non‐structured domains of the scaffold Nups and performed in vitro self‐assembled particle assays with those derived from channel‐forming FG‐Nups. We found that non‐structured domains of ELYS and Nup35N interacted with channel‐forming FG‐Nups to form a self‐assembled particle. Sequential addition of FG‐Nups into the scaffold particle revealed that ELYS, which initiates postmitotic NPC reassembly, interacts with early assembling FG‐Nups (Nups98 and 153) but not middle stage‐assembling FG‐Nups (Nups58 and 62). Nup35, which assembles between the early and middle stages, facilitated the assembly of Nup62 into the early assembling Nups both in vitro and in vivo. These results demonstrate that ELYS and Nup35 have a role of facilitator in the ordered assembly of channel‐forming FG‐Nups during mitosis. [ABSTRACT FROM AUTHOR]

Abstract Nuclear transport receptors (NTRs) recognize localization signals of cargos to facilitate their passage across the central channel of nuclear pore complexes (NPCs). About 30 different NTRs constitute different transport pathways in humans and bind to a multitude of different cargos. The exact cargo spectrum of the majority of NTRs, their specificity and even the extent to which active nucleocytoplasmic transport contributes to protein localization remains understudied because of the transient nature of these interactions and the wide dynamic range of cargo concentrations. To systematically map cargo–NTR relationships in situ, we used proximity ligation coupled to mass spectrometry (BioID). We systematically fused the engineered biotin ligase BirA* to 16 NTRs. We estimate that a considerable fraction of the human proteome is subject to active nuclear transport. We quantified the specificity and redundancy in NTR interactions and identified transport pathways for cargos. We extended the BioID method by the direct identification of biotinylation sites. This approach enabled us to identify interaction interfaces and to discriminate direct versus piggyback transport mechanisms. Data are available via ProteomeXchange with identifier PXD007976.

Background: LRP5/6 are co-receptors in Wnt/β-catenin pathway. Recently, we discovered multiple β-catenin independent functions of LRP5/6 in tumor cells and in the diseased heart. Nucleoporin 37 (NUP37) is an important component of the nuclear pore complex (NPC), whose elevated expression is associated with worsened prognosis in liver cancer. Previous studies have shown that NUP37 interacted with YAP and activated YAP/TEAD signaling in liver cancer. Our preliminary findings showed a nuclear location of LRP5. We thus tested the hypothesis that LRP5 may act as a genuine regulator of YAP/TEAD signaling via modulating NUP37 in a β-catenin-independent way. Methods: We performed siRNA knockdown of LRP5, LRP6, or β-catenin in liver cancer HepG2 cells to determine the effect on tumor cell proliferation. Protein expressions and interaction between LRP5 and NUP37 were determined using immunoprecipitation and western blot analyses. Results: HepG2 cell proliferation was markedly inhibited by knockdown of LRP5 but not LRP6 or β-catenin, suggesting that LRP5 has a specific, β-catenin-independent role in inhibiting HepG2 cell proliferation. Knockdown of NUP37 by siRNA inhibited the proliferation of HepG2 cells, whereas overexpression of NUP37 reversed the decrease in cell proliferation induced by LRP5 knockdown. Immunoprecipitation assays confirmed that LRP5 bound to NUP37. Furthermore, LRP5 overexpression restored NUP37 knockdown-induced downregulation of YAP/TEAD pathway. Conclusions: LRP5 deletion attenuates cell proliferation via destabilization of NUP37, in a β-catenin-independent manner. LRP5 therefore acts as a genuine regulator of YAP/TEAD signaling via maintaining the integrity of the NPC, and implicates a therapeutic strategy in targeting LRP5 for inhibiting liver cancer cell proliferation. [ABSTRACT FROM AUTHOR]

Introduction: The nuclear pore complex is comprised of approximately 30 proteins named nucleoporins (Nups) and tightly regulates nucleocytoplasmic transport of macromolecules across the nuclear membrane. Genetic alterations in many NUP genes are associated with many human maladies, such as neurological disease, autoimmune disorders and cancer. Methods: We reviewed the status quo of recent advancement of the knowledge of oncogenic role of nucleoporins in human carcinogenesis, focusing on major non-hematological malignancies in the recent literature. Both clinical study-derived and experiment-based reports were critically reviewed. We have also discussed the potential of nucleoporins as novel cancer biomarkers and promising therapeutic target against human malignancies. Results: Several Nups such as Nup53, Nup88, Nup98, Nup160 and Nup214 modulated a plethora of cellular and physiological pathways involved in tumorigenesis such as GSK3β-Snail, Wnt/β-Catenin and RanGap1/RanBP2 signaling axes, DNA damage response, resistance to apoptosis and chemotherapy. Conclusion: Although classically, majority of studies have shown oncogenic roles of nucleoporins as genetic fusion partners in several types of leukemia, emerging evidence suggests that nucleoporins also modulate many cellular signaling pathways that are associated with several major non-hematological malignancies, such as carcinomas of skin, breast, lung, prostate and colon. Hence, nucleoporins are emerging as novel therapeutic targets in human tumors. [ABSTRACT FROM AUTHOR]

Mutations or dysregulation of nucleoporins (Nups) are strongly associated with neural developmental diseases, yet the underlying mechanisms remain poorly understood. Here, we show that depletion of Nup Seh1 in radial glial progenitors results in defective neural progenitor proliferation and differentiation that ultimately manifests in impaired neurogenesis and microcephaly. This loss of stem cell proliferation is not associated with defects in the nucleocytoplasmic transport. Rather, transcriptome analysis showed that ablation of Seh1 in neural stem cells derepresses the expression of p21, and knockdown of p21 partially restored self-renewal capacity. Mechanistically, Seh1 cooperates with the NuRD transcription repressor complex at the nuclear periphery to regulate p21 expression. Together, these findings identified that Nups regulate brain development by exerting a chromatin-associated role and affecting neural stem cell proliferation. [Display omitted] • Nucleoporin Seh1 is essential for neurogenesis and neocortical development in mice • Seh1 cooperates with the NuRD transcription repressor complex • Seh1 regulates murine neural stem cell proliferation via silencing p21 expression Dai and Liu et al. demonstrate that nucleoporin Seh1 is essential for murine neural stem cell proliferation and differentiation. Seh1 cooperates with the NuRD transcription repressor complex to regulate p21 expression and defines a regulatory role of nucleoporin in the cell cycle and neurogenesis. [ABSTRACT FROM AUTHOR]

Nuclear organization has emerged as a critical layer in the coordination of DNA repair activities. Distinct types of DNA lesions have notably been shown to relocate at the vicinity of nuclear pore complexes (NPCs), where specific repair pathways are favored, ultimately safeguarding genome integrity. Here, we review the most recent progress in this field, notably highlighting the increasingly diverse types of DNA structures undergoing repositioning, and the signaling pathways involved. We further discuss our growing knowledge of the molecular mechanisms underlying the choice of repair pathways at NPCs, and their conservation — or divergences. Intriguingly, a series of recent findings suggest that DNA metabolism may be coupled to NPC biogenesis and specialization, challenging our initial vision of these processes. [ABSTRACT FROM AUTHOR]

Nuclear pore complexes (NPCs) mediate the bidirectional transport of cargo across the nuclear envelope (NE). NPCs are also membrane remodeling machines with a capacity to curve and fuse the membranes of the NE. However, little is known about the interplay of NPCs and lipids at a mechanistic level. A full understanding of NPC structure and function needs to encompass how the NPC shapes membranes and is itself shaped by lipids. Here we attempt to connect recent findings in NPC research with the broader field of membrane biochemistry to illustrate how an interplay between NPCs and lipids may facilitate the conformational plasticity of NPCs and the generation of a unique pore membrane topology. We highlight the need to better understand the NPC's lipid environment and outline experimental avenues towards that goal. [Display omitted] • Nuclear membranes shape nuclear pore architecture and diameter. • Balanced lipid saturation is critical for NPC integrity and function. • In cellulo structures suggest how a unique pore membrane topology is shaped. • Nucleoporin crowding on membranes could facilitate curvature generation. • Nuclear pores may be surrounded by a distinct lipid nano-environment. [ABSTRACT FROM AUTHOR]

At first glance the nucleus is a highly conserved organelle. Overall nuclear morphology, the octagonal nuclear pore complex, the presence of peripheral heterochromatin and the nuclear envelope appear near constant features right down to the ultrastructural level. New work is revealing significant compositional divergence within these nuclear structures and their associated functions, likely reflecting adaptations and distinct mechanisms between eukaryotic lineages and especially the trypanosomatids. While many examples of mechanistic divergence currently lack obvious functional interpretations, these studies underscore the malleability of nuclear architecture. I will discuss some recent findings highlighting these facets within trypanosomes, together with the underlying evolutionary framework and make a call for the exploration of nuclear function in non-canonical experimental organisms. [ABSTRACT FROM AUTHOR]

The delivery of the HIV-1 genome into the nucleus is an indispensable step in retroviral infection of non-dividing cells, but the mechanism of HIV-1 nuclear import has been a longstanding debate due to controversial experimental evidence. It was commonly believed that the HIV-1 capsid would need to disassemble (uncoat) in the cytosol before nuclear import because the capsid is larger than the central channel of nuclear pore complexes (NPCs); however, increasing evidence demonstrates that intact, or nearly intact, HIV-1 capsid passes through the NPC to enter the nucleus. With the protection of the capsid, the HIV-1 core completes reverse transcription in the nucleus and is translocated to the integration site. Uncoating occurs while, or after, the viral genome is released near the integration site. These independent discoveries reveal a compelling new paradigm of this important step of the HIV-1 life cycle. In this review, we summarize the recent studies related to HIV-1 nuclear import, highlighting the spatial–temporal relationship between the nuclear entry of the virus core, reverse transcription, and capsid uncoating.

Specific macromolecules are rapidly transported across the nuclear envelope via the nuclear pore complex (NPC). The selective transport process is facilitated when nuclear transport receptors (NTRs) weakly and transiently bind to intrinsically disordered constituents of the NPC, FG Nups. These two types of proteins help maintain the selective NPC barrier. To interrogate their binding interactions in vitro, we deployed an NPC barrier mimic. We created the stationary phase by covalently attaching fragments of a yeast FG Nup called Nsp1 to glass coverslips. We used a tunable mobile phase containing NTR, nuclear transport factor 2 (NTF2). In the stationary phase, three main factors affected binding: the number of FG repeats, the charge of fragments, and the fragment density. We also identified three main factors affecting binding in the mobile phase: the avidity of the NTF2 variant for Nsp1, the presence of nonspecific proteins, and the presence of additional NTRs. We used both experimentally determined binding parameters and molecular dynamics simulations of Nsp1FG fragments to create an agent-based model. The results suggest that NTF2 binding is negatively cooperative and dependent on the density of Nsp1FG molecules. Our results demonstrate the strengths of combining experimental and physical modeling approaches to study NPC-mediated transport.

Cellular stress induces the formation of membraneless protein condensates in both the nucleus and cytoplasm. The nucleocytoplasmic transport of proteins mainly occurs through nuclear pore complexes (NPCs), whose efficiency is affected by various stress conditions. Here, we report that hyperosmotic stress compartmentalizes nuclear 26S proteasomes into dense nuclear foci, independent of signaling cascades. Most of the proteasome foci were detected between the condensed chromatin mass and inner nuclear membrane. The proteasome-positive puncta were not colocalized with other types of nuclear bodies and were reversibly dispersed when cells were returned to the isotonic medium. The structural integrity of 26S proteasomes in the nucleus was slightly affected under the hyperosmotic condition. We also found that these insulator-body-like proteasome foci were possibly formed through disrupted nucleus-to-cytosol transport, which was mediated by the sequestration of NPC components into osmostress-responding stress granules. These data suggest that phase separation in both the nucleus and cytosol may be a major cell survival mechanism during hyperosmotic stress conditions.

The name “eukaryote” is derived from Greek, meaning “true kernel”, and describes the domain of organisms whose cells have a nucleus. The nucleus is thus the defining feature of eukaryotes and distinguishes them from prokaryotes (Archaea and Bacteria), whose cells lack nuclei. Despite this, we discuss the intriguing possibility that organisms on the path from the first eukaryotic common ancestor to the last common ancestor of all eukaryotes did not possess a nucleus at all—at least not in a form we would recognize today—and that the nucleus in fact arrived relatively late in the evolution of eukaryotes. The clues to this alternative evolutionary path lie, most of all, in recent discoveries concerning the structure of the nuclear pore complex. We discuss the evidence for such a possibility and how this impacts our views of eukaryote origins and how eukaryotes have diversified subsequent to their last common ancestor.

Nuclear pore complexes (NPCs) at the surface of nuclear membranes play a critical role in regulating the transport of both small molecules and macromolecules between the cell nucleus and cytoplasm via their multilayered spiderweb-like central channel. During mitosis, nuclear envelope breakdown leads to the rapid disintegration of NPCs, allowing some NPC proteins to play crucial roles in the kinetochore structure, spindle bipolarity, and centrosome homeostasis. The aberrant functioning of nucleoporins (Nups) and NPCs has been associated with autoimmune diseases, viral infections, neurological diseases, cardiomyopathies, and cancers, especially leukemia. This Special Issue highlights several new contributions to the understanding of NPC proteostasis.

Transport from and into the nucleus is essential to all eukaryotic life and occurs through the nuclear pore complex (NPC). There are a multitude of data supporting a role for nuclear transport in neurodegenerative diseases, but actual transport assays in disease models have provided diverse outcomes. In this review, we summarize how nuclear transport works, which transport assays are available, and what matters complicate the interpretation of their results. Taking a specific type of ALS caused by mutations in C9orf72 as an example, we illustrate these complications, and discuss how the current data do not firmly answer whether the kinetics of nucleocytoplasmic transport are altered. Answering this open question has far-reaching implications, because a positive answer would imply that widespread mislocalization of proteins occurs, far beyond the reported mislocalization of transport reporters, and specific proteins such as FUS, or TDP43, and thus presents a challenge for future research.

The nuclear basket (NB) scaffold, a fibrillar structure anchored to the nuclear pore complex (NPC), is regarded as constructed of polypeptides of the coiled-coil dominated protein TPR to which other proteins can bind without contributing to the NB’s structural integrity. Here we report vertebrate protein ZC3HC1 as a novel inherent constituent of the NB, common at the nuclear envelopes (NE) of proliferating and non-dividing, terminally differentiated cells of different morphogenetic origin. Formerly described as a protein of other functions, we instead present the NB component ZC3HC1 as a protein required for enabling distinct amounts of TPR to occur NB-appended, with such ZC3HC1-dependency applying to about half the total amount of TPR at the NEs of different somatic cell types. Furthermore, pointing to an NB structure more complex than previously anticipated, we discuss how ZC3HC1 and the ZC3HC1-dependent TPR polypeptides could enlarge the NB’s functional repertoire.

Picornaviruses are positive-stranded RNA viruses. Even though replication and translation of their genome take place in the cytoplasm, these viruses evolved different strategies to disturb nucleocytoplasmic trafficking of host proteins and RNA. The major targets of picornavirus are the phenylalanine-glycine (FG)-nucleoporins, which form a mesh in the central channel of the nuclear pore complex through which protein cargos and karyopherins are actively transported in both directions. Interestingly, while enteroviruses use the proteolytic activity of their 2A protein to degrade FG-nucleoporins, cardioviruses act by triggering phosphorylation of these proteins by cellular kinases. By targeting the nuclear pore complex, picornaviruses recruit nuclear proteins to the cytoplasm, where they increase viral genome translation and replication; they affect nuclear translocation of cytoplasmic proteins such as transcription factors that induce innate immune responses and retain host mRNA in the nucleus thereby preventing cell emergency responses and likely making the ribosomal machinery available for translation of viral RNAs.

The nucleoporins Nup60p, Nup2p, and Nup1p form part of the nuclear basket structure of the Saccharomyces cerevisiae nuclear pore complex (NPC). Here, we show that these necleoporins can be isolated from yeast extracts by affinity chromatography on karyopherin Kap95p-coated beads. To characterize Nup60p further, Nup60p-coated beads were used to capture its interacting proteins from extracts. We find that Nup60p binds to Nup2p and serves as a docking site for Kap95p-Kap60p heterodimers and Kap123p. Nup60p also binds Gsp1p-GTP and its guanine nucleotide exchange factor Prp20p, and functions as a Gsp1p guanine nucleotide dissociation inhibitor by reducing the activity of Prp20p. Yeast lacking Nup60p exhibit minor defects in nuclear export of Kap60p, nuclear import of Kap95p-Kap60p-dependent cargoes, and diffusion of small proteins across the NPC. Yeast lacking Nup60p also fail to anchor Nup2p at the NPC, resulting in the mislocalization of Nup2p to the nucleoplasm and cytoplasm. Purified Nup60p and Nup2p bind each other directly, but the stability of the complex is compromised when Kap60p binds Nup2p. Gsp1p-GTP enhances by 10-fold the affinity between Nup60p and Nup2p, and restores binding of Nup2p-Kap60p complexes to Nup60p. The results suggest a dynamic interaction, controlled by the nucleoplasmic concentration of Gsp1p-GTP, between Nup60p and Nup2p at the NPC.

Production of healthy gametes in meiosis relies on the quality control and proper distribution of both nuclear and cytoplasmic contents. Meiotic differentiation naturally eliminates age-induced cellular damage by an unknown mechanism. Using time-lapse fluorescence microscopy in budding yeast, we found that nuclear senescence factors – including protein aggregates, extrachromosomal ribosomal DNA circles, and abnormal nucleolar material – are sequestered away from chromosomes during meiosis II and subsequently eliminated. A similar sequestration and elimination process occurs for the core subunits of the nuclear pore complex in both young and aged cells. Nuclear envelope remodeling drives the formation of a membranous compartment containing the sequestered material. Importantly, de novo generation of plasma membrane is required for the sequestration event, preventing the inheritance of long-lived nucleoporins and senescence factors into the newly formed gametes. Our study uncovers a new mechanism of nuclear quality control and provides insight into its function in meiotic cellular rejuvenation.