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EMBO Reports Dec 2023Protein translocation across the endoplasmic reticulum (ER) membrane is an essential step during protein entry into the secretory pathway. The conserved Sec61...
Protein translocation across the endoplasmic reticulum (ER) membrane is an essential step during protein entry into the secretory pathway. The conserved Sec61 protein-conducting channel facilitates polypeptide translocation and coordinates cotranslational polypeptide-processing events. In cells, the majority of Sec61 is stably associated with a heterotetrameric membrane protein complex, the translocon-associated protein complex (TRAP), yet the mechanism by which TRAP assists in polypeptide translocation remains unknown. Here, we present the structure of the core Sec61/TRAP complex bound to a mammalian ribosome by cryogenic electron microscopy (cryo-EM). Ribosome interactions anchor the Sec61/TRAP complex in a conformation that renders the ER membrane locally thinner by significantly curving its lumenal leaflet. We propose that TRAP stabilizes the ribosome exit tunnel to assist nascent polypeptide insertion through Sec61 and provides a ratcheting mechanism into the ER lumen mediated by direct polypeptide interactions.
Topics: Animals; SEC Translocation Channels; Membrane Proteins; Endoplasmic Reticulum; Mammals; Peptides; Protein Transport
PubMed: 37983950
DOI: 10.15252/embr.202357910 -
Nature Communications Aug 2023Extracellular vesicles (EVs) are gaining ground as next-generation drug delivery modalities. Genetic fusion of the protein of interest to a scaffold protein with high...
Extracellular vesicles (EVs) are gaining ground as next-generation drug delivery modalities. Genetic fusion of the protein of interest to a scaffold protein with high EV-sorting ability represents a robust cargo loading strategy. To address the paucity of such scaffold proteins, we leverage a simple and reliable assay that can distinguish intravesicular cargo proteins from surface- as well as non-vesicular proteins and compare the EV-sorting potential of 244 candidate proteins. We identify 24 proteins with conserved EV-sorting abilities across five types of producer cells. TSPAN2 and TSPAN3 emerge as lead candidates and outperform the well-studied CD63 scaffold. Importantly, these engineered EVs show promise as delivery vehicles in cell cultures and mice as demonstrated by efficient transfer of luminal cargo proteins as well as surface display of different functional entities. The discovery of these scaffolds provides a platform for EV-based engineering.
Topics: Mice; Animals; Extracellular Vesicles; Proteins; Drug Delivery Systems; Protein Transport; Cell Communication
PubMed: 37550290
DOI: 10.1038/s41467-023-40453-0 -
The Chinese Journal of Dental Research Mar 2024Coordination and information exchange among the various organelles ensure the precise and orderly functioning of eukaryotic cells. Interaction between the cytoplasm and... (Review)
Review
Coordination and information exchange among the various organelles ensure the precise and orderly functioning of eukaryotic cells. Interaction between the cytoplasm and nucleoplasm is crucial for many physiological processes. Macromolecular protein transport into the nucleus requires assistance from the nuclear transport system. These proteins typically contain a nuclear localisation sequence that guides them to enter the nucleus. Understanding the mechanism of nuclear import of macromolecular proteins is important for comprehending cellular processes. Investigation of disease-related alterations can facilitate the development of novel therapeutic strategies and provide additional evidence for clinical trials. This review provides an overview of the proteins involved in nuclear transport and the mechanisms underlying macromolecular protein transport.
Topics: Active Transport, Cell Nucleus; Protein Transport; Cell Nucleus; Cytoplasm; Eukaryotic Cells
PubMed: 38546518
DOI: 10.3290/j.cjdr.b5136729 -
Traffic (Copenhagen, Denmark) Sep 2023Endosomal cargo recycling lies at the heart of subcellular trafficking processes under the management of several Ras-related GTP-binding proteins (Rabs) which are... (Review)
Review
Endosomal cargo recycling lies at the heart of subcellular trafficking processes under the management of several Ras-related GTP-binding proteins (Rabs) which are coordinated by their upstream regulators and require their downstream effectors to display their functions. In this regard, several Rabs have been well-reviewed except Rab22a. Rab22a is a crucial regulator of vesicle trafficking, early endosome and recycling endosome formation. Notably, recent studies demonstrated the immunological roles of Rab22a, which are closely associated with cancers, infection and autoimmune disorders. This review provides an overview of the regulators and effectors of Rab22a. Also, we highlight the current knowledge of the role of Rab22a in endosomal cargo recycling, including the biogenesis of recycling tubules with the help of a complex with Rab22a at its core, and how different internalized cargo chooses different recycling routes thanks to the cooperation of Rab22a, its effectors and its regulators. Of note, contradictions and speculation related to endosomal cargo recycling that Rab22a brings impacts on are also discussed. Finally, this review endeavors to briefly introduce the various events impacted by Rab22a, particularly focusing on the commandeered Rab22a-associated endosomal maturation and endosomal cargo recycling, in addition to the extensively investigated oncogenic role of Rab22a.
Topics: Protein Transport; rab GTP-Binding Proteins; Endosomes; Cell Communication
PubMed: 37340959
DOI: 10.1111/tra.12907 -
Nature Chemical Biology Sep 2023The Sec61 complex forms a protein-conducting channel in the endoplasmic reticulum membrane that is required for secretion of soluble proteins and production of many...
The Sec61 complex forms a protein-conducting channel in the endoplasmic reticulum membrane that is required for secretion of soluble proteins and production of many membrane proteins. Several natural and synthetic small molecules specifically inhibit Sec61, generating cellular effects that are useful for therapeutic purposes, but their inhibitory mechanisms remain unclear. Here we present near-atomic-resolution structures of human Sec61 inhibited by a comprehensive panel of structurally distinct small molecules-cotransin, decatransin, apratoxin, ipomoeassin, mycolactone, cyclotriazadisulfonamide and eeyarestatin. All inhibitors bind to a common lipid-exposed pocket formed by the partially open lateral gate and plug domain of Sec61. Mutations conferring resistance to the inhibitors are clustered at this binding pocket. The structures indicate that Sec61 inhibitors stabilize the plug domain in a closed state, thereby preventing the protein-translocation pore from opening. Our study provides the atomic details of Sec61-inhibitor interactions and the structural framework for further pharmacological studies and drug design.
Topics: Humans; Endoplasmic Reticulum; Membrane Proteins; Protein Transport; SEC Translocation Channels
PubMed: 37169959
DOI: 10.1038/s41589-023-01337-y -
Cells Dec 2023Explaining changes at the gene level that occur during neurodegeneration in the CA3 area is crucial from the point of view of memory impairment and the development of...
Explaining changes at the gene level that occur during neurodegeneration in the CA3 area is crucial from the point of view of memory impairment and the development of post-ischemic dementia. An ischemic model of Alzheimer's disease was used to evaluate changes in the expression of genes related to amyloid transport in the CA3 region of the hippocampus after 10 min of brain ischemia with survival of 2, 7 and 30 days and 12, 18 and 24 months. The quantitative reverse transcriptase PCR assay revealed that the expression of the and genes involved in amyloid transport was dysregulated from 2 days to 24 months post-ischemia in the CA3 area of the hippocampus. gene expression 2 and 7 days after ischemia was below control values. However, its expression from day 30 to 24 months, survival after an ischemic episode was above control values. gene expression 2 days after ischemia was below control values, reaching a maximum increase 7 and 30 days post-ischemia. Then, after 12, 18 and 24 months, it was again below the control values. The data indicate that in the CA3 area of the hippocampus, an episode of brain ischemia causes the increased expression of the gene for 7-30 days during the acute phase and that of from 1 to 24 months after ischemia during the chronic stage. In other words, in the early post-ischemic stage, the expression of the gene that transport amyloid to the brain increases (7-30 days). Conversely, in the late post-ischemic stage, amyloid scavenging/cleaning gene activity increases, reducing and/or preventing further neuronal damage or facilitating the healing of damaged sites. This is how the new phenomenon of pyramidal neuronal damage in the CA3 area after ischemia is defined. In summary, post-ischemic modification of the and genes is useful in the study of the ischemic pathways and molecular factors involved in the development of Alzheimer's disease.
Topics: Humans; Alzheimer Disease; Amyloidogenic Proteins; Brain Ischemia; Hippocampus; Ischemia; Low Density Lipoprotein Receptor-Related Protein-1; tau Proteins; Protein Transport
PubMed: 38067191
DOI: 10.3390/cells12232763 -
The New Phytologist Oct 2023The endomembrane system consists of various membrane-bound organelles including the endoplasmic reticulum (ER), Golgi apparatus, trans-Golgi network (TGN), endosomes,... (Review)
Review
The endomembrane system consists of various membrane-bound organelles including the endoplasmic reticulum (ER), Golgi apparatus, trans-Golgi network (TGN), endosomes, and the lysosome/vacuole. Membrane trafficking between distinct compartments is mainly achieved by vesicular transport. As the endomembrane compartments and the machineries regulating the membrane trafficking are largely conserved across all eukaryotes, our current knowledge on organelle biogenesis and endomembrane trafficking in plants has mainly been shaped by corresponding studies in mammals and yeast. However, unique perspectives have emerged from plant cell biology research through the characterization of plant-specific regulators as well as the development and application of the state-of-the-art microscopical techniques. In this review, we summarize our current knowledge on the plant endomembrane system, with a focus on several distinct pathways: ER-to-Golgi transport, protein sorting at the TGN, endosomal sorting on multivesicular bodies, vacuolar trafficking/vacuole biogenesis, and the autophagy pathway. We also give an update on advanced imaging techniques for the plant cell biology research.
Topics: Plants; Endosomes; Vacuoles; Multivesicular Bodies; Protein Transport; Golgi Apparatus; trans-Golgi Network
PubMed: 37507353
DOI: 10.1111/nph.19134 -
Experimental Dermatology Nov 2023PKM2 mediates the Warburg effects and is crucial for tumorigenesis, but its role in hyperplastic skin disorders remains elusive. In this study, we investigated the...
PKM2 mediates the Warburg effects and is crucial for tumorigenesis, but its role in hyperplastic skin disorders remains elusive. In this study, we investigated the function of PKM2 in psoriatic keratinocytes. We found that PKM2 expression and its nuclear translocation were induced in the epidermis of psoriasis patients, contributing to aerobic glycolysis and cell growth. Moreover, mass spectrometry combined with immunoprecipitation analysis revealed that PKM2 could interact with TRIM33, an E3 ubiquitin ligase in the nucleus, and this interaction is critical for the nuclear retention of PKM2. As a result of TRIM33-mediated ubiquitination, PKM2 nuclear protein kinase function is promoted, thus leading to the phosphorylation of STAT3. In addition, blocking PKM2 nuclear translocation abrogated TRIM33-triggered glycolysis and cell proliferation in keratinocytes. Taken together, our experiments demonstrate that ubiquitination regulates the nuclear retention of PKM2 in keratinocytes. Moreover, our results highlight a novel mechanism accounting for the metabolic reprogramming of keratinocytes in psoriasis patients.
Topics: Humans; Cell Line, Tumor; Glycolysis; Keratinocytes; Phosphorylation; Protein Transport; Psoriasis; Transcription Factors; Thyroid Hormone-Binding Proteins
PubMed: 37688280
DOI: 10.1111/exd.14922 -
The New Phytologist Aug 2023Secretion is a fundamental process that plant pathogens utilize to deliver effectors into the host to downregulate immunity and promote infection. Here, we uncover a...
Secretion is a fundamental process that plant pathogens utilize to deliver effectors into the host to downregulate immunity and promote infection. Here, we uncover a fascinating membrane trafficking and delivery route that originates from vacuolar membranes in Magnaporthe oryzae and conduits to the host interface and plasma membrane. To perform such secretory/trafficking function, MoRab7 first recruits the retromer complex to the vacuolar membrane, enabling recognition of a family of SNARE proteins, including MoSnc1. Live-cell imaging confirmed a highly dynamic vesicular trafficking of the retromer complex component(s) and MoSnc1 toward and across the host interface or plasma membrane, and subsequent fusion with target membranes. Interestingly, disruption of the MoRab7/Retromer/MoSnc1-based endolysosomal cascade affects effector secretion and fungal pathogenicity. Taken together, we discovered an unconventional protein and membrane trafficking route starting from the fungal endolysosomes to the M. oryzae-rice interaction interface and dissect the role of MoRab7/Retromer/MoSnc1 sorting machinery in effector secretion during biotrophy and invasive growth in rice blast fungus.
Topics: Endosomes; Protein Transport; Vacuoles; Biological Transport; Cell Membrane; Magnaporthe; Oryza; Fungal Proteins; Plant Diseases
PubMed: 37291895
DOI: 10.1111/nph.19050 -
Nature Communications Oct 2023Cytosolic metalloenzymes acquire metals from buffered intracellular pools. How exported metalloenzymes are appropriately metalated is less clear. We provide evidence...
Cytosolic metalloenzymes acquire metals from buffered intracellular pools. How exported metalloenzymes are appropriately metalated is less clear. We provide evidence that TerC family proteins function in metalation of enzymes during export through the general secretion (Sec-dependent) pathway. Bacillus subtilis strains lacking MeeF(YceF) and MeeY(YkoY) have a reduced capacity for protein export and a greatly reduced level of manganese (Mn) in the secreted proteome. MeeF and MeeY copurify with proteins of the general secretory pathway, and in their absence the FtsH membrane protease is essential for viability. MeeF and MeeY are also required for efficient function of the Mn-dependent lipoteichoic acid synthase (LtaS), a membrane-localized enzyme with an extracytoplasmic active site. Thus, MeeF and MeeY, representative of the widely conserved TerC family of membrane transporters, function in the co-translocational metalation of Mn-dependent membrane and extracellular enzymes.
Topics: Bacterial Proteins; Protein Transport; Bacillus subtilis; Secretory Pathway; Metalloproteins
PubMed: 37794032
DOI: 10.1038/s41467-023-41896-1