-
The Journal of Cell Biology Apr 2023ER tubules form and maintain membrane contact sites (MCSs) with late endosomes/lysosomes (LE/lys). The molecular composition and cellular functions of these MCSs are...
ER tubules form and maintain membrane contact sites (MCSs) with late endosomes/lysosomes (LE/lys). The molecular composition and cellular functions of these MCSs are poorly understood. Here, we find that Tex2, an SMP domain-containing lipid transfer protein conserved in metazoan and yeast, is a tubular ER protein and is recruited to ER-LE/lys MCSs by TMEM55, phosphatases that convert PI(4,5)P2 to PI5P on LE/lys. We show that the Tex2-TMEM55 interaction occurs between an N-terminal region of Tex2 and a catalytic motif in the PTase domain of TMEM55. The Tex2-TMEM55 interaction can be regulated by endosome-resident type 2 PI4K activities. Functionally, Tex2 knockout results in defects in lysosomal trafficking, digestive capacity, and lipid composition of LE/lys membranes. Together, our data identify Tex2 as a tubular ER protein that resides at TMEM55-dependent ER-LE/lys MCSs required for lysosomal functions.
Topics: Animals; Endoplasmic Reticulum; Endosomes; Lysosomes; Carrier Proteins
PubMed: 36705603
DOI: 10.1083/jcb.202205133 -
Nature Communications Jul 2020Endosomes are compositionally dynamic organelles that regulate signaling, nutrient status and organelle quality by specifying whether material entering the cells will be...
Endosomes are compositionally dynamic organelles that regulate signaling, nutrient status and organelle quality by specifying whether material entering the cells will be shuttled back to the cell surface or degraded by the lysosome. Recently, membrane contact sites (MCSs) between the endoplasmic reticulum (ER) and endosomes have emerged as important players in endosomal protein sorting, dynamics and motility. Here, we show that PDZD8, a Synaptotagmin-like Mitochondrial lipid-binding Proteins (SMP) domain-containing ER transmembrane protein, utilizes distinct domains to interact with Rab7-GTP and the ER transmembrane protein Protrudin and together these components localize to an ER-late endosome MCS. At these ER-late endosome MCSs, mitochondria are also recruited to form a three-way contact. Thus, our data indicate that PDZD8 is a shared component of two distinct MCSs and suggest a role for SMP-mediated lipid transport in the regulation of endosome function.
Topics: Adaptor Proteins, Signal Transducing; Biological Transport; Cell Line; Endoplasmic Reticulum; Endosomes; Humans; Membrane Proteins; Mitochondria; Mitochondrial Proteins; Protein Transport; Vesicular Transport Proteins; rab GTP-Binding Proteins; rab7 GTP-Binding Proteins
PubMed: 32686675
DOI: 10.1038/s41467-020-17451-7 -
ELife Nov 2021Cell-cell communication is an essential process in life, with endosomes acting as key organelles for regulating uptake and secretion of signaling molecules. Endocytosed...
Cell-cell communication is an essential process in life, with endosomes acting as key organelles for regulating uptake and secretion of signaling molecules. Endocytosed material is accepted by the sorting endosome where it either is sorted for recycling or remains in the endosome as it matures to be degraded in the lysosome. Investigation of the endosome maturation process has been hampered by the small size and rapid movement of endosomes in most cellular systems. Here, we report an easy versatile live-cell imaging assay to monitor endosome maturation kinetics, which can be applied to a variety of mammalian cell types. Acute ionophore treatment led to enlarged early endosomal compartments that matured into late endosomes and fused with lysosomes to form endolysosomes. Rab5-to-Rab7 conversion and PI(3)P formation and turn over were recapitulated with this assay and could be observed with a standard widefield microscope. We used this approach to show that Snx1 and Rab11-positive recycling endosome recruitment occurred throughout endosome maturation and was uncoupled from Rab conversion. In contrast, efficient endosomal acidification was dependent on Rab conversion. The assay provides a powerful tool to further unravel various aspects of endosome maturation.
Topics: Endosomes; HeLa Cells; Humans; Lysosomes; Microscopy, Fluorescence
PubMed: 34846303
DOI: 10.7554/eLife.70982 -
Traffic (Copenhagen, Denmark) Sep 2001Late endosomes, which have the morphological characteristics of multivesicular bodies, have received relatively little attention in comparison with early endosomes and... (Review)
Review
Late endosomes, which have the morphological characteristics of multivesicular bodies, have received relatively little attention in comparison with early endosomes and lysosomes. Recent work in mammalian and yeast cells has given insights into their structure and function, including the generation of their multivesicular morphology. Lipid partitioning to create microdomains enriched in specific lipids is observed in late endosomes, with some lumenal vesicles enriched in lysobisphosphatidic acid and others in phosphatidylinositol 3-phosphate. Sorting of membrane proteins into the lumenal vesicles may occur because of the properties of their trans-membrane domains, or as a result of tagging with ubiquitin. Yeast class E Vps proteins and their mammalian orthologs are the best candidates to make up the protein machinery that controls inward budding, a process that starts in early endosomes. Late endosomes are able to undergo homotypic fusion events and also heterotypic fusion with lysosomes, a process that delivers endocytosed macromolecules for proteolytic degradation.
Topics: Animals; Endocytosis; Endosomes; Lipid Metabolism; Lysosomes; Models, Biological; Phosphatidylinositol Phosphates; Protein Structure, Tertiary; Protein Transport
PubMed: 11555415
DOI: 10.1034/j.1600-0854.2001.20904.x -
EMBO Reports Mar 2021The endoplasmic reticulum (ER) is the largest organelle contacting virtually every other organelle for information exchange and control of processes such as transport,...
The endoplasmic reticulum (ER) is the largest organelle contacting virtually every other organelle for information exchange and control of processes such as transport, fusion, and fission. Here, we studied the role of the other organelles on ER network architecture in the cell periphery. We show that the co-migration of the ER with other organelles, called ER hitchhiking facilitated by late endosomes and lysosomes is a major mechanism controlling ER network architecture. When hitchhiking occurs, emerging ER structures may fuse with the existing ER tubules to alter the local ER architecture. This couples late endosomal/lysosomal positioning and mobility to ER network architecture. Conditions restricting late endosomal movement-including cell starvation-or the depletion of tether proteins that link the ER to late endosomes reduce ER dynamics and limit the complexity of the peripheral ER network architecture. This indicates that among many factors, the ER is controlled by late endosomal movement resulting in an alteration of the ER network architecture.
Topics: Biological Transport; Endoplasmic Reticulum; Endosomes; Lysosomes
PubMed: 33554435
DOI: 10.15252/embr.202050815 -
Journal of Cell Science Apr 2021Rab5 and Rab7a are the main determinants of early and late endosomes and are important regulators of endosomal progression. The transport from early endosomes to late...
Rab5 and Rab7a are the main determinants of early and late endosomes and are important regulators of endosomal progression. The transport from early endosomes to late endosome seems to be regulated through an endosomal maturation switch, where Rab5 is gradually exchanged by Rab7a on the same endosome. Here, we provide new insight into the mechanism of endosomal maturation, for which we have discovered a stepwise Rab5 detachment, sequentially regulated by Rab7a. The initial detachment of Rab5 is Rab7a independent and demonstrates a diffusion-like first-phase exchange between the cytosol and the endosomal membrane, and a second phase, in which Rab5 converges into specific domains that detach as a Rab5 indigenous endosome. Consequently, we show that early endosomal maturation regulated through the Rab5-to-Rab7a switch induces the formation of new fully functional Rab5-positive early endosomes. Progression through stepwise early endosomal maturation regulates the direction of transport and, concomitantly, the homeostasis of early endosomes.
Topics: Endosomes; Transport Vesicles; rab5 GTP-Binding Proteins
PubMed: 33737317
DOI: 10.1242/jcs.254185 -
ACS Applied Materials & Interfaces Jul 2022Lipid nanoparticles (LNPs) are important delivery systems for RNA-based therapeutics, yet the mechanism of their interaction with endosomal membranes remains unclear....
Lipid nanoparticles (LNPs) are important delivery systems for RNA-based therapeutics, yet the mechanism of their interaction with endosomal membranes remains unclear. Here, the interactions of nucleic acid-loaded LNPs that contain an ionizable lipid with models of the early and late endosomal membranes are studied, for the first time, using different reflectometry techniques. Novel insight is provided with respect to the subphase pH, the stage of the endosome, and the nature of the nucleic acid cargo. It is found that the insertion of lipids from the LNPs into the model membrane is greatest at pH 6.5 and 5.5, whereas at higher pH, lipid insertion is suppressed with evidence instead for the binding of intact LNPs, demonstrating the importance of the pH in the fusion of LNPs undergoing the endosomal pathway. Furthermore, and independently of the pH, the effect of the early- versus late-stage endosomal models is minimal, suggesting that the increased fluidity and anionic nature of the late endosome has little effect on the extent of LNP interaction. Last, there is greater nucleic acid delivery from LNPs containing mRNA than Poly(A), indicating that the extent of interaction can be tuned according to the nature of the nucleic acid cargo. Such new information on the relative impact of factors influencing nucleic acid delivery by LNP interactions with endosomal membranes is important in the design and tuning of vehicles with improved nucleic acid delivery capacities.
Topics: Endosomes; Lipids; Liposomes; Nanoparticles; RNA, Messenger; RNA, Small Interfering
PubMed: 35758331
DOI: 10.1021/acsami.2c06065 -
The Journal of Neuroscience : the... Nov 2021, which encodes the protein tSNARE1, is a high-confidence gene candidate for schizophrenia risk, but nothing is known about its cellular or physiological function. We...
, which encodes the protein tSNARE1, is a high-confidence gene candidate for schizophrenia risk, but nothing is known about its cellular or physiological function. We identified the major gene products of and their cytoplasmic localization and function in endosomal trafficking in cortical neurons. We validated three primary isoforms of expressed in human brain, all of which encode a syntaxin-like Qa SNARE domain. RNA-sequencing data from adult and fetal human brain suggested that the majority of tSNARE1 lacks a transmembrane domain that is thought to be necessary for membrane fusion. Biochemical data demonstrate that tSNARE1 can compete with Stx12 for incorporation into an endosomal SNARE complex, supporting its possible role as an inhibitory SNARE. Live-cell imaging in cortical neurons from mice of both sexes demonstrated that brain tSNARE1 isoforms localized to the endosomal network. The most abundant brain isoform, tSNARE1c, localized most frequently to Rab7 late endosomes, and endogenous tSNARE1 displayed a similar localization in human neural progenitor cells and neuroblastoma cells. In mature rat neurons from both sexes, tSNARE1 localized to the dendritic shaft and dendritic spines, supporting a role for tSNARE1 at the postsynapse. Expression of either tSNARE1b or tSNARE1c, which differ only in their inclusion or exclusion of an Myb-like domain, delayed the trafficking of the dendritic endosomal cargo Nsg1 into late endosomal and lysosomal compartments. These data suggest that tSNARE1 regulates endosomal trafficking in cortical neurons, likely by negatively regulating early endosomal to late endosomal trafficking. Schizophrenia is a severe and polygenic neuropsychiatric disorder. Understanding the functions of high-confidence candidate genes is critical toward understanding how their dysfunction contributes to schizophrenia pathogenesis. is one of the high-confidence candidate genes for schizophrenia risk, yet nothing was known about its cellular or physiological function. Here we describe the major isoforms of and their cytoplasmic localization and function in the endosomal network in cortical neurons. Our results are consistent with the hypothesis that the majority of brain tSNARE1 acts as a negative regulator to endolysosomal trafficking.
Topics: Animals; Cerebral Cortex; Endosomes; Female; Humans; Male; Mice; Mice, Inbred C57BL; Neurons; Protein Isoforms; Protein Transport; Rats; Rats, Sprague-Dawley; SNARE Proteins; Schizophrenia
PubMed: 34642214
DOI: 10.1523/JNEUROSCI.0556-21.2021 -
Developmental Cell Jan 2018The endomembrane system consists of the secretory and endocytic pathways, which communicate by transport to and from the trans-Golgi network (TGN). In mammalian cells,...
The endomembrane system consists of the secretory and endocytic pathways, which communicate by transport to and from the trans-Golgi network (TGN). In mammalian cells, the endocytic pathway includes early, late, and recycling endosomes. In budding yeast, different types of endosomes have been described, but the organization of the endocytic pathway has remained unclear. We performed a spatial and temporal analysis of yeast endosomal markers and endocytic cargoes. Our results indicate that the yeast TGN also serves as an early and recycling endosome. In addition, as previously described, yeast contains a late or prevacuolar endosome (PVE). Endocytic cargoes localize to the TGN shortly after internalization, and manipulations that perturb export from the TGN can slow the passage of endocytic cargoes to the PVE. Yeast apparently lacks a distinct early endosome. Thus, yeast has a simple endocytic pathway that may reflect the ancestral organization of the endomembrane system.
Topics: Biological Transport; Cell Membrane; Endocytosis; Endosomes; Golgi Apparatus; Saccharomycetales; trans-Golgi Network
PubMed: 29316441
DOI: 10.1016/j.devcel.2017.12.014 -
Journal of Biochemistry Oct 2020Inositol phospholipids are low-abundance regulatory lipids that orchestrate diverse cellular functions in eukaryotic organisms. Recent studies have uncovered involvement... (Review)
Review
Inositol phospholipids are low-abundance regulatory lipids that orchestrate diverse cellular functions in eukaryotic organisms. Recent studies have uncovered involvement of the lipids in multiple steps in autophagy. The late endosome-lysosome compartment plays critical roles in cellular nutrient sensing and in the control of both the initiation of autophagy and the late stage of eventual degradation of cytosolic materials destined for elimination. It is particularly notable that inositol lipids are involved in almost all steps of the autophagic process. In this review, we summarize how inositol lipids regulate and contribute to autophagy through the endomembrane compartments, primarily focusing on PI4P and PI(4,5)P2.
Topics: Animals; Autophagy; Endosomes; Humans; Lysosomes; Phosphatidylinositol 4,5-Diphosphate; Phosphatidylinositol Phosphates; Phosphatidylinositols; Signal Transduction
PubMed: 32745205
DOI: 10.1093/jb/mvaa089