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The EMBO Journal Jul 2023Nucleoporins (Nups) assemble nuclear pores that form the permeability barrier between nucleoplasm and cytoplasm. Nucleoporins also localize in cytoplasmic foci proposed...
Nucleoporins (Nups) assemble nuclear pores that form the permeability barrier between nucleoplasm and cytoplasm. Nucleoporins also localize in cytoplasmic foci proposed to function as pore pre-assembly intermediates. Here, we characterize the composition and incidence of cytoplasmic Nup foci in an intact animal, C. elegans. We find that, in young non-stressed animals, Nup foci only appear in developing sperm, oocytes and embryos, tissues that express high levels of nucleoporins. The foci are condensates of highly cohesive FG repeat-containing nucleoporins (FG-Nups), which are maintained near their solubility limit in the cytoplasm by posttranslational modifications and chaperone activity. Only a minor fraction of FG-Nup molecules concentrate in Nup foci, which dissolve during M phase and are dispensable for nuclear pore assembly. Nucleoporin condensation is enhanced by stress and advancing age, and overexpression of a single FG-Nup in post-mitotic neurons is sufficient to induce ectopic condensation and organismal paralysis. We speculate that Nup foci are non-essential and potentially toxic condensates whose assembly is actively suppressed in healthy cells.
Topics: Male; Animals; Nuclear Pore Complex Proteins; Nuclear Pore; Caenorhabditis elegans; Semen; Cell Nucleus; Active Transport, Cell Nucleus
PubMed: 37254647
DOI: 10.15252/embj.2022112987 -
Nature Communications Oct 2023Receptor-mediated transport of soluble proteins is nature's key to empowering eukaryotic cells to access a plethora of macromolecules, either by direct accumulation or...
Receptor-mediated transport of soluble proteins is nature's key to empowering eukaryotic cells to access a plethora of macromolecules, either by direct accumulation or as products from resulting biochemical pathways. The transport efficiency of these mechanisms results from the receptor's capability to capture, transport, and release ligands on the one hand and the cycling ability that allows for performing multiple rounds of ligand transport on the other. However, the plant VACUOLAR SORTING RECEPTOR (VSR) protein family is diverse, and their ligand-specificity and bidirectional trafficking routes and transport mechanisms remain highly controversial. Here we employ nanobody-epitope interaction-based molecular tools to assess the function of the VSR 7 in vivo. We demonstrate the specificity of the VSR7 for sequence-specific vacuolar sorting signals, and we trace its anterograde transport and retrograde recycling route. VSR7 localizes at the cis-Golgi apparatus at steady state conditions and transports ligands downstream to release them in the trans-Golgi network/early endosome (TGN/EE) before undergoing clathrin-dependent recycling from the TGN/EE back to the cis-Golgi.
Topics: trans-Golgi Network; Clathrin; Ligands; Golgi Apparatus; Protein Transport; Carrier Proteins; Plant Proteins; Endosomes
PubMed: 37903761
DOI: 10.1038/s41467-023-42331-1 -
Cell Reports Jan 2024Fanconi anemia (FA) is characterized by congenital abnormalities, bone marrow failure, and cancer susceptibility. The central FA protein complex FANCI/FANCD2 (ID2) is...
Fanconi anemia (FA) is characterized by congenital abnormalities, bone marrow failure, and cancer susceptibility. The central FA protein complex FANCI/FANCD2 (ID2) is activated by monoubiquitination and recruits DNA repair proteins for interstrand crosslink (ICL) repair and replication fork protection. Defects in the FA pathway lead to R-loop accumulation, which contributes to genomic instability. Here, we report that the splicing factor SRSF1 and FANCD2 interact physically and act together to suppress R-loop formation via mRNA export regulation. We show that SRSF1 stimulates FANCD2 monoubiquitination in an RNA-dependent fashion. In turn, FANCD2 monoubiquitination proves crucial for the assembly of the SRSF1-NXF1 nuclear export complex and mRNA export. Importantly, several SRSF1 cancer-associated mutants fail to interact with FANCD2, leading to inefficient FANCD2 monoubiquitination, decreased mRNA export, and R-loop accumulation. We propose a model wherein SRSF1 and FANCD2 interaction links DNA damage response to the avoidance of pathogenic R-loops via regulation of mRNA export.
Topics: Humans; R-Loop Structures; Active Transport, Cell Nucleus; Fanconi Anemia; Fanconi Anemia Complementation Group Proteins; Fanconi Anemia Complementation Group D2 Protein; Ubiquitination; DNA Repair; Neoplasms; RNA, Messenger; DNA Damage; Serine-Arginine Splicing Factors
PubMed: 38165804
DOI: 10.1016/j.celrep.2023.113610 -
Physiological Research Apr 2024This comprehensive review explores the physiological and pathophysiological significance of VPS13A, a protein encoded by the VPS13A gene. The VPS13A gene is associated... (Review)
Review
This comprehensive review explores the physiological and pathophysiological significance of VPS13A, a protein encoded by the VPS13A gene. The VPS13A gene is associated with Chorea-acanthocytosis (ChAc), a rare hereditary neurodegenerative disorder. The review covers essential aspects, beginning with the genetics of VPS13A, highlighting its role in the pathogenesis of ChAc, and addressing the spectrum of genetic variants involved. It delves into the structure and function of the VPS13A protein, emphasizing its presence in various tissues and its potential involvement in protein trafficking and lipid homeostasis. Molecular functions of VPS13A in the brain tissue and other cell types or tissues with respect to their role in cytoskeletal regulation and autophagy are explored. Finally, it explores the intriguing link between VPS13A mutations, lipid imbalances, and neurodegeneration, shedding light on future research directions. Overall, this review serves as a comprehensive resource for understanding the pivotal role of VPS13A in health and disease, particularly in the context of ChAc. Key words: Chorein , Tumor, Actin, Microfilament, Gene expression, Chorea-acanthocytosis.
Topics: Humans; Animals; Vesicular Transport Proteins; Neuroacanthocytosis; Mutation; Lipid Metabolism
PubMed: 38710051
DOI: 10.33549/physiolres.935268 -
Trends in Cell Biology May 2024Peroxisomes are vital metabolic organelles that import their lumenal (matrix) enzymes from the cytosol using mobile receptors. Surprisingly, the receptors can even... (Review)
Review
Peroxisomes are vital metabolic organelles that import their lumenal (matrix) enzymes from the cytosol using mobile receptors. Surprisingly, the receptors can even import folded proteins, but the underlying mechanism has been a mystery. Recent results reveal how import receptors shuttle cargo into peroxisomes. The cargo-bound receptors move from the cytosol across the peroxisomal membrane completely into the matrix by a mechanism that resembles transport through the nuclear pore. The receptors then return to the cytosol through a separate retrotranslocation channel, leaving the cargo inside the organelle. This cycle concentrates imported proteins within peroxisomes, and the energy for cargo import is supplied by receptor export. Peroxisomal protein import thus fundamentally differs from other previously known mechanisms for translocating proteins across membranes.
Topics: Peroxisomes; Protein Transport; Humans; Animals; Cytosol; Receptors, Cytoplasmic and Nuclear
PubMed: 37743160
DOI: 10.1016/j.tcb.2023.08.005 -
Cell Chemical Biology Aug 2023Insulin resistance (IR) is the root cause of type II diabetes, yet no safe treatment is available to address it. Using a high throughput compatible assay that measures...
Insulin resistance (IR) is the root cause of type II diabetes, yet no safe treatment is available to address it. Using a high throughput compatible assay that measures real-time translocation of the glucose transporter glucose transporter 4 (GLUT4), we identified small molecules that potentiate insulin action. In vivo, these insulin sensitizers improve insulin-stimulated GLUT4 translocation, glucose tolerance, and glucose uptake in a model of IR. Using proteomic and CRISPR-based approaches, we identified the targets of those compounds as Unc119 proteins and solved the structure of Unc119 bound to the insulin sensitizer. This study identifies compounds that have the potential to be developed into diabetes treatment and establishes Unc119 proteins as targets for improving insulin sensitivity.
Topics: Humans; Insulin; Diabetes Mellitus, Type 2; Proteomics; Glucose; Protein Transport; Insulin Resistance; Glucose Transport Proteins, Facilitative; Glucose Transporter Type 4
PubMed: 37453421
DOI: 10.1016/j.chembiol.2023.06.012 -
Journal of Cell Science Oct 2023The malaria-causing parasite, Plasmodium falciparum completely remodels its host red blood cell (RBC) through the export of several hundred parasite proteins, including...
The malaria-causing parasite, Plasmodium falciparum completely remodels its host red blood cell (RBC) through the export of several hundred parasite proteins, including transmembrane proteins, across multiple membranes to the RBC. However, the process by which these exported membrane proteins are extracted from the parasite plasma membrane for export remains unknown. To address this question, we fused the exported membrane protein, skeleton binding protein 1 (SBP1), with TurboID, a rapid, efficient and promiscuous biotin ligase (SBP1TbID). Using time-resolved proximity biotinylation and label-free quantitative proteomics, we identified two groups of SBP1TbID interactors - early interactors (pre-export) and late interactors (post-export). Notably, two promising membrane-associated proteins were identified as pre-export interactors, one of which possesses a predicted translocon domain, that could facilitate the export of membrane proteins. Further investigation using conditional mutants of these candidate proteins showed that these proteins were essential for asexual growth and localize to the host-parasite interface during early stages of the intraerythrocytic cycle. These data suggest that they might play a role in ushering membrane proteins from the parasite plasma membrane for export to the host RBC.
Topics: Animals; Humans; Biotinylation; Erythrocytes; Malaria; Plasmodium falciparum; Porins; Protein Transport; Protozoan Proteins
PubMed: 37772444
DOI: 10.1242/jcs.260506 -
Sheng Wu Gong Cheng Xue Bao = Chinese... Nov 2023Yeast surface display (YSD) is a technology that fuses the exogenous target protein gene sequence with a specific vector gene sequence, followed by introduction into... (Review)
Review
Yeast surface display (YSD) is a technology that fuses the exogenous target protein gene sequence with a specific vector gene sequence, followed by introduction into yeast cells. Subsequently, the target protein is expressed and localized on the yeast cell surface by using the intracellular protein transport mechanism of yeast cells, whereas the most widely used YSD system is the α-agglutinin expression system. Yeast cells possess the eukaryotic post-translational modification mechanism, which helps the target protein fold correctly. This mechanism could be used to display various eukaryotic proteins, including antibodies, receptors, enzymes, and antigenic peptides. YSD has become a powerful protein engineering tool in biotechnology and biomedicine, and has been used to improve a broad range of protein properties including affinity, specificity, enzymatic function, and stability. This review summarized recent advances in the application of YSD technology from the aspects of library construction and screening, antibody engineering, protein engineering, enzyme engineering and vaccine development.
Topics: Saccharomyces cerevisiae; Protein Engineering; Biotechnology; Antibodies; Amino Acid Sequence
PubMed: 38013172
DOI: 10.13345/j.cjb.230085 -
The Journal of Biological Chemistry Nov 2023T-cell receptor stimulation triggers cytosolic Ca signaling by inositol-1,4,5-trisphosphate (IP)-mediated Ca release from the endoplasmic reticulum (ER) and Ca entry...
T-cell receptor stimulation triggers cytosolic Ca signaling by inositol-1,4,5-trisphosphate (IP)-mediated Ca release from the endoplasmic reticulum (ER) and Ca entry through Ca release-activated Ca (CRAC) channels gated by ER-located stromal-interacting molecules (STIM1/2). Physiologically, cytosolic Ca signaling manifests as regenerative Ca oscillations, which are critical for nuclear factor of activated T-cells-mediated transcription. In most cells, Ca oscillations are thought to originate from IP receptor-mediated Ca release, with CRAC channels indirectly sustaining them through ER refilling. Here, experimental and computational evidence support a multiple-oscillator mechanism in Jurkat T-cells whereby both IP receptor and CRAC channel activities oscillate and directly fuel antigen-evoked Ca oscillations, with the CRAC channel being the major contributor. KO of either STIM1 or STIM2 significantly reduces CRAC channel activity. As such, STIM1 and STIM2 synergize for optimal Ca oscillations and activation of nuclear factor of activated T-cells 1 and are essential for ER refilling. The loss of both STIM proteins abrogates CRAC channel activity, drastically reduces ER Ca content, severely hampers cell proliferation and enhances cell death. These results clarify the mechanism and the contribution of STIM proteins to Ca oscillations in T-cells.
Topics: Humans; Calcium; Calcium Release Activated Calcium Channels; Calcium Signaling; Jurkat Cells; Stromal Interaction Molecule 1; Stromal Interaction Molecule 2; Gene Knockout Techniques; Models, Biological; Protein Isoforms; Protein Transport; Cell Proliferation; Cell Survival
PubMed: 37778728
DOI: 10.1016/j.jbc.2023.105310 -
International Journal of Molecular... Sep 2023Breast cancer represents a paramount global health challenge, warranting intensified exploration of the molecular underpinnings influencing its progression to facilitate... (Review)
Review
Breast cancer represents a paramount global health challenge, warranting intensified exploration of the molecular underpinnings influencing its progression to facilitate the development of precise diagnostic instruments and customized therapeutic regimens. Historically, the Golgi apparatus has been acknowledged for its primary role in protein sorting and trafficking within cellular contexts. However, recent findings suggest a potential link between modifications in Golgi apparatus function and organization and the pathogenesis of breast cancer. This review delivers an exhaustive analysis of this correlation. Specifically, we examine the consequences of disrupted protein glycosylation, compromised protein transport, and inappropriate oncoprotein processing on breast cancer cell dynamics. Furthermore, we delve into the impacts of Golgi-mediated secretory routes on the release of pro-tumorigenic factors during the course of breast cancer evolution. Elucidating the nuanced interplay between the Golgi apparatus and breast cancer can pave the way for innovative therapeutic interventions and the discovery of biomarkers, potentially enhancing the diagnostic, prognostic, and therapeutic paradigms for afflicted patients. The advancement of such research could substantially expedite the realization of these objectives.
Topics: Humans; Female; Breast Neoplasms; Breast; Golgi Apparatus; Carcinogenesis; Biological Transport
PubMed: 37762375
DOI: 10.3390/ijms241814073