NUP62 Human

What is NUP62 and its primary function in human cells?

NUP62 (Nucleoporin p62) is an essential component of the nuclear pore complex (NPC), which forms a versatile transport channel between the nucleus and cytoplasm. The NPC is a highly modular assembly consisting of approximately 34 distinct nucleoporins (Nups), with NUP62 playing a critical role in nucleocytoplasmic transport . NUP62 contains phenylalanine-glycine (FG) repeats that contribute to the selective permeability barrier of the NPC, allowing specific proteins to traverse between nuclear and cytoplasmic compartments . In addition to its canonical role in nuclear transport, emerging evidence suggests NUP62 functions beyond the nuclear pore, including in centrosome homeostasis and cell division .

How is NUP62 organized within the nuclear pore complex architecture?

NUP62 is present in multiple subcomplexes of the nuclear pore complex, including the central transport channel (CTC) and the cytoplasmic ring (CR) . Structural characterization has revealed that the NUP62 coiled-coil domain (amino acids 322-525 in mammals) is critical for its interactions with other nucleoporins, particularly Nup88 and Nup214 . When reconstituted in vitro, the CR complex containing NUP62 forms a distinct "4"-shaped architecture that resembles the CTC complex . NUP62 is often found in a subcomplex with O-glycosylated proteins of molecular masses 62, 58, 54, and 45 kDa, forming a core component that lines the central channel of the nuclear pore .

What are the known protein interactions of NUP62 that mediate its function?

NUP62 participates in multiple protein-protein interactions that facilitate its diverse functions:

• NPC components: NUP62 interacts with Nup88 (amino acids 517-742) and Nup214 (amino acids 693-926) to form heterotrimers that are essential for NPC assembly .

• Transport mediators: NUP62 binds to Nup93 (amino acids 1-150), which helps anchor it within the NPC structure .

• Heat shock proteins: NUP62 interacts with hsp90, hsp70, p23, and TPR domain proteins FKBP52 and PP5 during nuclear import processes .

• Exocyst complex: NUP62 binds the N-terminal domain of the exocyst component Exo70 through its coiled-coil domain, suggesting a role in cellular transport beyond the nuclear envelope .

• Centrosomal proteins: NUP62 interacts with gamma-tubulin and SAS-6, which are critical components of centrosomes, indicating its role in centrosome homeostasis .

What role does NUP62 play in cell division and centrosome function?

NUP62 maintains centrosome integrity through several mechanisms, with its depletion causing profound effects on cell division. Research has demonstrated that knockdown of NUP62 induces mitotic arrest in G2/M phases and subsequent mitotic cell death . Depletion of NUP62 through RNA interference results in:

• Defective centrosome segregation and centriole maturation during G2 phase

• Formation of multipolar centrosomes

• Centriole synthesis defects

• Dramatic spindle orientation defects

• Rearrangement of centrosome components that impair cell bipolarity

Evidence suggests NUP62 is crucial for targeting gamma-tubulin and SAS-6 to the centrioles, explaining its significant impact on centrosome function and cellular division . The appearance of multinucleated cells following NUP62 depletion further emphasizes its importance in maintaining genomic stability through proper centrosome function.

How does NUP62 expression change during cellular stress responses?

Chronic stress significantly impacts NUP62 expression and localization. In chronologically stressed animals, particularly within the hippocampal CA3 region, there is a selective reduction in translated Nup62 transcripts . This reduction appears to occur through two distinct mechanisms:

• Reduced translation: Genetically targeted translating ribosome affinity purification revealed a selective reduction in translated Nup62 transcripts in CA3 neurons of chronically stressed mice .

• Post-translational modification: Phosphorylation of NUP62 at a specific tyrosine residue (Y422 in humans) is associated with its shedding from the nuclear pore complex and/or retention in the cytoplasm .

The stress-induced phosphorylation appears to be mediated by proline-rich tyrosine kinase 2 (PYK2), which shows subcellular redistribution in chronically stressed pyramidal neurons . This stress-related modulation of NUP62 may contribute to dendritic shrinkage in hippocampal neurons, providing a molecular link between chronic stress and morphological changes in neurons.

What is the relationship between NUP62 and neurological function or disease?

NUP62 has significant implications for neurological function and disease through multiple mechanisms:

• Dendritic morphology: Diminishing NUP62 content in cultured hippocampal neurons results in simplification and shortening of dendritic arbors, which may contribute to stress-induced cognitive impairment and depression .

• Basal ganglia degeneration: Mutations in NUP62 cause autosomal recessive infantile bilateral striatal necrosis, indicating its crucial role in basal ganglia development and maintenance .

• Neurodegenerative diseases: NUP62 belongs to the nucleoporin family, several members of which have been implicated in neurodegenerative conditions such as frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), and Parkinson's disease (PD) . For example, motor neuron-specific knockout of Nup358, another nucleoporin, causes an ALS-like phenotype in mice .

These connections highlight the importance of proper nucleocytoplasmic transport, mediated in part by NUP62, for neuronal health and function.

What are the most effective approaches for studying NUP62 localization and dynamics in living cells?

Researchers studying NUP62 localization and dynamics can employ several complementary techniques:

• Fluorescent protein tagging: Creating NUP62-GFP (or other fluorescent protein) fusions for live-cell imaging to track its movement and localization. This approach should target domains that don't interfere with NUP62's function, typically avoiding the central FG-repeat domain.

• Photoactivatable or photoconvertible tags: Using proteins like PA-GFP or Dendra2 fused to NUP62 allows for pulse-chase experiments to track the movement and half-life of NUP62 populations in living cells.

• Fluorescence recovery after photobleaching (FRAP): This technique can assess the dynamic association of NUP62 with the nuclear pore complex by bleaching a region and measuring recovery time.

• Super-resolution microscopy: Techniques such as STORM or PALM provide nanoscale resolution to visualize NUP62 within the NPC architecture, revealing its precise localization relative to other nucleoporins.

• Proximity labeling: Methods like BioID or APEX2 fused to NUP62 can identify proteins in close proximity to NUP62 in living cells, revealing its interaction network in different cellular compartments.

When designing these experiments, researchers should consider that phosphorylation of NUP62 at Y422 (human) may affect its localization, potentially causing shedding from the NPC or cytoplasmic retention .

What techniques are recommended for effectively depleting NUP62 in experimental systems?

Several approaches have been successfully employed to deplete NUP62 in experimental systems:

• RNA interference (RNAi): siRNA or shRNA targeting NUP62 has been effectively used to deplete the protein, resulting in observable phenotypes including defective centrosome segregation and mitotic arrest . When designing RNAi experiments, include proper controls to account for off-target effects.

• CRISPR-Cas9 gene editing: For complete knockout or endogenous tagging of NUP62. Complete knockout may not be viable in many cell types, so inducible systems or partial knockouts might be necessary.

• Degron-based approaches: Fusion of inducible degron tags to endogenous NUP62 allows for rapid, conditional protein depletion, which can be useful for studying acute effects without compensatory mechanisms.

• Dominant negative approaches: Expression of truncated versions of NUP62 (particularly the coiled-coil domain) can disrupt interactions with partner proteins like Nup88 and Nup214, interfering with proper NPC assembly .

When depleting NUP62, researchers should be aware that complete knockout may lead to cell death due to its essential functions in nucleocytoplasmic transport and mitosis . Therefore, partial depletion or time-controlled depletion systems are often more informative.

How can researchers effectively study NUP62 phosphorylation and its functional consequences?

Studying NUP62 phosphorylation requires specialized techniques to detect and manipulate this post-translational modification:

• Phospho-specific antibodies: Use antibodies specifically targeting phosphorylated Y422 (human) to detect this modification by western blotting or immunofluorescence .

• Mass spectrometry: Phosphoproteomic analysis can identify all phosphorylation sites on NUP62, including those that may not be previously characterized.

• Phosphomimetic and phosphodeficient mutants: Creating Y422E (phosphomimetic) or Y422F (phosphodeficient) mutations allows for studying the functional consequences of phosphorylation at this site.

• Kinase inhibitors: PYK2 inhibitors can be used to determine if preventing phosphorylation of NUP62 at Y422 rescues stress-induced phenotypes .

• Subcellular fractionation: This technique can separate nuclear and cytoplasmic fractions to quantify the distribution of phosphorylated NUP62, which has been shown to accumulate in cytoplasmic fractions under stress conditions .

When studying phosphorylation, it's important to consider that stress conditions, particularly chronic stress, may influence the phosphorylation status of NUP62 and its subsequent localization and function .

How does NUP62 contribute to the biomolecular condensation properties of the nuclear pore complex?

NUP62 contains intrinsically disordered FG-repeat domains that contribute to the phase separation properties of the nuclear pore complex central channel. This distinctive feature is critical for understanding selective transport:

• FG-repeat hydrogel formation: Nsp1, the yeast homolog of Nup62, has been used to form in vitro phenylalanine-glycine (FG)-repeat hydrogels that mimic fundamental aspects of selective protein trafficking . This suggests that the core biophysical features enabling selective transport through the NPC are encoded within the amino acid sequence of NUP62.

• Phase separation properties: The FG-repeats in NUP62 can undergo liquid-liquid phase separation, creating a selective permeability barrier that allows passage of transport receptors while excluding other macromolecules.

• Interaction with transport receptors: Transport receptors like importins and exportins can partition into these FG-repeat condensates, enabling their passage through the central channel along with their cargoes.

Researchers investigating these properties should consider techniques such as in vitro reconstitution of purified NUP62 domains, fluorescence correlation spectroscopy to measure partitioning of molecules into NUP62 condensates, and microrheology to characterize the material properties of these biomolecular condensates.

What is the crosstalk between NUP62 and chromatin organization?

Beyond its role in nucleocytoplasmic transport, NUP62 functions in chromatin organization and gene expression regulation:

• Chromatin interactions: NUP62 has been implicated in functions related to chromatin organization, particularly at the nuclear periphery . These interactions may influence gene expression patterns and nuclear architecture.

• Gene expression regulation: Changes in NUP62 levels or function can impact transcriptional programs. For example, in the prefrontal cortex of major depressives, NUP62 transcripts are decreased, suggesting a potential role in the gene expression changes associated with depression .

• Epigenetic regulation: NPCs, including components like NUP62, can create microenvironments at the nuclear periphery that influence the epigenetic status of associated chromatin regions.

To study these interactions, researchers can employ techniques such as ChIP-seq to identify genomic regions associated with NUP62, DamID for mapping nuclear periphery interactions, and RNA-seq following NUP62 depletion to identify affected gene expression programs.

How does the developmental and tissue-specific expression of NUP62 influence its function in different cellular contexts?

NUP62 expression and function varies across development and different tissue contexts:

• Developmental regulation: While not explicitly detailed in the search results, other nucleoporins show critical developmental functions, with many nucleoporin null mutations resulting in embryonic lethality . This suggests that NUP62 may also have developmentally regulated functions.

• Tissue-specific expression patterns: NUP62 may have tissue-specific expression levels or isoforms that contribute to specialized functions in different cell types. For example, its role in neuronal cells may differ from its function in dividing cells.

• Conditional phenotypes: In plants, overexpression-based co-suppression of AtNup62 leads to severely dwarfed, early flowering plants, suggesting important functions in plant development . Similar conditional phenotypes may exist in human tissues.

Understanding these tissue-specific and developmental roles requires techniques such as single-cell RNA-seq to map expression patterns, tissue-specific conditional knockout models, and comparative proteomics to identify tissue-specific interaction partners of NUP62.

What potential therapeutic targets exist within the NUP62 pathway for neurological conditions?

Several aspects of NUP62 biology present potential therapeutic targets for neurological conditions:

• PYK2 inhibition: Given that proline-rich tyrosine kinase 2 (PYK2) appears to mediate stress-induced phosphorylation of NUP62, PYK2 inhibitors could potentially prevent the disassociation of NUP62 from the NPC and subsequent dendritic shrinkage in chronic stress conditions .

• NUP62 stabilization: Approaches that stabilize NUP62 levels or prevent its stress-induced reduction could potentially mitigate the neuronal changes associated with chronic stress and depression.

• NPC transport modulation: As NUP62 is critical for nucleocytoplasmic transport, compounds that selectively enhance this function might compensate for reduced NUP62 levels in conditions like stress-induced depression.

• Targeting NUP62 in infantile bilateral striatal necrosis: Since mutations in NUP62 cause autosomal recessive infantile bilateral striatal necrosis, gene therapy approaches to restore functional NUP62 in affected individuals could be explored .

These therapeutic approaches would require careful development to ensure specificity and minimize disruption of essential cellular functions dependent on NUP62.

How does altered NUP62 function contribute to the pathophysiology of stress-related disorders and neurodegeneration?

The pathophysiological contributions of NUP62 dysfunction include:

• Dendritic atrophy: Chronic stress reduces NUP62 levels in hippocampal CA3 neurons, and diminished NUP62 content results in simplification and shortening of dendritic arbors . This structural change may underlie cognitive impairments associated with chronic stress and depression.

• Nucleocytoplasmic transport disruption: Alterations in NUP62 function could disrupt the proper transport of proteins and RNA between the nucleus and cytoplasm, affecting numerous cellular processes including gene expression and stress responses.

• Centrosome dysfunction: Given NUP62's role in centrosome integrity, disruption could lead to abnormal cell division in neural progenitors or impact neuronal migration during development .

• Connection to neurodegenerative mechanisms: Other nucleoporins have been implicated in neurodegenerative conditions like frontotemporal dementia, amyotrophic lateral sclerosis, and Parkinson's disease . Similar mechanisms may apply to NUP62 dysfunction.

Understanding these pathophysiological mechanisms provides insights into potential therapeutic interventions for stress-related disorders and neurodegenerative conditions involving NUP62 dysfunction.

What are the key domains and post-translational modifications of human NUP62?

Table 1: Key Domains and Modifications of Human NUP62

The coiled-coil domain (amino acids 322-525) is particularly critical for forming heterotrimers with other nucleoporins including Nup88 and Nup214, which are essential components of the nuclear pore complex .

What phenotypes are associated with NUP62 and related nucleoporin mutations?

Table 2: Phenotypes Associated with Nucleoporin Mutations

This table illustrates the range of phenotypes associated with nucleoporin mutations, from embryonic lethality to specific disease states, highlighting their essential roles in cellular function.

What experimental systems and models are most suitable for studying NUP62 function?

Table 3: Experimental Systems for NUP62 Research

Each experimental system offers distinct advantages for investigating different aspects of NUP62 biology, and researchers should select the most appropriate system based on their specific research questions.

What are the most promising areas for future research on NUP62 function?

Several promising research directions for NUP62 include:

• Structural biology approaches: Cryo-electron microscopy and X-ray crystallography studies of NUP62 in complex with its interacting partners to understand the molecular details of NPC assembly and function.

• Single-molecule tracking: Advanced imaging techniques to track individual NUP62 molecules within living cells to understand its dynamics and transport mechanisms.

• Tissue-specific functions: Investigation of NUP62's role in different tissues, particularly in the nervous system where mutations lead to specific pathologies.

• Stress response mechanisms: Further elucidation of how chronic stress impacts NUP62 function and localization, and whether these changes are reversible.

• Therapeutic targeting: Development of approaches to modulate NUP62 function or prevent its stress-induced depletion for potential applications in stress-related disorders.

These research directions would significantly advance our understanding of NUP62 biology and its implications for human health and disease.

How might emerging technologies advance our understanding of NUP62 biology?

Emerging technologies that could transform NUP62 research include:

• Cryo-electron tomography: This technique can visualize NPCs in their native cellular environment at near-atomic resolution, providing insights into the structural organization of NUP62 within the intact NPC.

• Proximity proteomics: Methods like BioID, APEX, and TurboID can map the protein interaction network of NUP62 in different cellular compartments and under various conditions.

• Single-cell multi-omics: Combining transcriptomics, proteomics, and epigenomics at the single-cell level can reveal how NUP62 function varies across cell types and states.

• Optical control techniques: Optogenetic approaches to manipulate NUP62 function with spatiotemporal precision could reveal its dynamic roles in cellular processes.

• Artificial intelligence for image analysis: Advanced machine learning algorithms can extract subtle patterns from imaging data, potentially revealing new aspects of NUP62 localization and dynamics.