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Nature Communications May 2024Tools for accessing and studying organelles remain underdeveloped. Here, we present a method by which giant organelle vesicles (GOVs) are generated by submitting cells...
Tools for accessing and studying organelles remain underdeveloped. Here, we present a method by which giant organelle vesicles (GOVs) are generated by submitting cells to a hypotonic medium followed by plasma membrane breakage. By this means, GOVs ranging from 3 to over 10 µm become available for micromanipulation. GOVs are made from organelles such as the endoplasmic reticulum, endosomes, lysosomes and mitochondria, or in contact with one another such as giant mitochondria-associated ER membrane vesicles. We measure the mechanical properties of each organelle-derived GOV and find that they have distinct properties. In GOVs procured from Cos7 cells, for example, bending rigidities tend to increase from the endoplasmic reticulum to the plasma membrane. We also found that the mechanical properties of giant endoplasmic reticulum vesicles (GERVs) vary depending on their interactions with other organelles or the metabolic state of the cell. Lastly, we demonstrate GERVs' biochemical activity through their capacity to synthesize triglycerides and assemble lipid droplets. These findings underscore the potential of GOVs as valuable tools for studying the biophysics and biology of organelles.
Topics: Animals; Chlorocebus aethiops; COS Cells; Endoplasmic Reticulum; Intracellular Membranes; Cell Membrane; Mitochondria; Organelles; Lipid Droplets; Triglycerides; Humans; Lysosomes
PubMed: 38704407
DOI: 10.1038/s41467-024-48086-7 -
Bioarchitecture 2016The early/recycling endosomes of an eukaryotic cell perform diverse cellular functions. In addition, the endosomal system generates multiple organelles, including... (Review)
Review
The early/recycling endosomes of an eukaryotic cell perform diverse cellular functions. In addition, the endosomal system generates multiple organelles, including certain cell type-specific organelles called lysosome-related organelles (LROs). The biosynthesis of these organelles possibly occurs through a sequential maturation process in which the cargo-containing endosomal vesicular/tubular structures are fused with the maturing organelle. The molecular machinery that regulates the cargo delivery or the membrane fusion during LRO biogenesis is poorly understood. Here, we describe the known key molecules, such as SNAREs, that regulate both the biogenesis and secretion of multiple LROs. Moreover, we also describe other regulatory molecules, such as Rab GTPases and their effectors that modulate the SNARE activity for cargo delivery to one such LRO, the melanosome. Overall, this review will increase our current understanding of LRO biogenesis and function.
Topics: Animals; Humans; Lysosomes; Organelles; SNARE Proteins
PubMed: 26760525
DOI: 10.1080/19490992.2015.1131890 -
Current Opinion in Cell Biology Apr 2015Maintaining the proper lipid composition of cellular membranes is critical for numerous cellular processes but mechanisms of membrane lipid homeostasis are not well... (Review)
Review
Maintaining the proper lipid composition of cellular membranes is critical for numerous cellular processes but mechanisms of membrane lipid homeostasis are not well understood. There is growing evidence that membrane contact sites (MCSs), regions where two organelles come in close proximity to one another, play major roles in the regulation of intracellular lipid composition and distribution. MCSs are thought to mediate the exchange of lipids and signals between organelles. In this review, we discuss how lipid exchange occurs at MCSs and evidence for roles of MCSs in regulating lipid synthesis and degradation. We also discuss how networks of organelles connected by MCSs may modulate cellular lipid homeostasis and help determine organelle lipid composition.
Topics: Binding Sites; Homeostasis; Intracellular Membranes; Lipid Metabolism; Mitochondria; Organelles
PubMed: 25569848
DOI: 10.1016/j.ceb.2014.12.004 -
Current Opinion in Cell Biology Aug 2023Membrane contact sites (MCSs) are areas of close proximity between organelles, implicated in transport of small molecules and in organelle biogenesis. Lipid transfer... (Review)
Review
Membrane contact sites (MCSs) are areas of close proximity between organelles, implicated in transport of small molecules and in organelle biogenesis. Lipid transfer proteins at MCSs facilitate the distribution of lipid species between organelle membranes. Such exchange processes rely on the apposition of two different membranes delimiting distinct compartments and a cytosolic intermembrane space. Maintaining organelle identity while transferring molecules therefore implies control over MCS architecture both on the ultrastructural and molecular levels. Factors including intermembrane distance, density of resident proteins, and contact surface area fine-tune MCS function. Furthermore, the structural arrangement of lipid transfer proteins and associated proteins underpins the molecular mechanisms of lipid fluxes at MCSs. Thus, the architecture of MCSs emerges as an essential aspect of their function.
Topics: Organelles; Mitochondrial Membranes; Mitochondria; Lipid Metabolism; Lipids
PubMed: 37515839
DOI: 10.1016/j.ceb.2023.102212 -
BMC Biology Jan 2018Caenorhabditis elegans neurons have recently been found to throw out cellular debris for remote degradation and/or storage, adding an "extracellular garbage elimination"... (Review)
Review
Caenorhabditis elegans neurons have recently been found to throw out cellular debris for remote degradation and/or storage, adding an "extracellular garbage elimination" option to known intracellular protein and organelle degradation pathways. This Q&A describes initial insights into the biology of seemingly selective protein and organelle elimination by challenged neurons, highlighting mysteries of how garbage is distinguished and sorted in the sending neuron, how the garbage-filled "exophers" appear to elicit degradative responses as they transit neighboring tissue, and how non-digestible materials get thrown out of cells again via processes that may be highly relevant to human neurodegenerative disease mechanisms.
Topics: Animals; Humans; Neurons; Organelles; Protein Transport; Proteolysis
PubMed: 29382333
DOI: 10.1186/s12915-018-0487-6 -
Current Biology : CB Jul 2014A tendency in cell biology is to divide and conquer. For example, decades of painstaking work have led to an understanding of endoplasmic reticulum (ER) and Golgi... (Review)
Review
A tendency in cell biology is to divide and conquer. For example, decades of painstaking work have led to an understanding of endoplasmic reticulum (ER) and Golgi structure, dynamics, and transport. In parallel, cytoskeletal researchers have revealed a fantastic diversity of structure and cellular function in both actin and microtubules. Increasingly, these areas overlap, necessitating an understanding of both organelle and cytoskeletal biology. This review addresses connections between the actin/microtubule cytoskeletons and organelles in animal cells, focusing on three key areas: ER structure and function; ER-to-Golgi transport; and Golgi structure and function. Making these connections has been challenging for several reasons: the small sizes and dynamic characteristics of some components; the fact that organelle-specific cytoskeletal elements can easily be obscured by more abundant cytoskeletal structures; and the difficulties in imaging membranes and cytoskeleton simultaneously, especially at the ultrastructural level. One major concept is that the cytoskeleton is frequently used to generate force for membrane movement, with two potential consequences: translocation of the organelle, or deformation of the organelle membrane. While initially discussing issues common to metazoan cells in general, we subsequently highlight specific features of neurons, since these highly polarized cells present unique challenges for organellar distribution and dynamics.
Topics: Actin Cytoskeleton; Animals; Endoplasmic Reticulum; Golgi Apparatus; Humans; Mice; Microtubules; Neurons; Organelles
PubMed: 25050967
DOI: 10.1016/j.cub.2014.05.033 -
Current Opinion in Cell Biology Apr 2016Peroxisomes participate in lipid metabolism, and are a major source of ROS in the cell. Their importance in cellular energy balance and redox homeostasis is... (Review)
Review
Peroxisomes participate in lipid metabolism, and are a major source of ROS in the cell. Their importance in cellular energy balance and redox homeostasis is well-established, as is the need to maintain peroxisome homeostasis to prevent pathologies associated with too few, or too many, of these organelles. How cells regulate peroxisome number has remained somewhat elusive. Recently, the tumor suppressors ATM and TSC, which regulate mTORC1 signaling, have been localized to peroxisomes. When activated by peroxisomal ROS, ATM signals to TSC to repress mTORC1 signaling and increase autophagic flux in cells, and also phosphorylates the peroxisomal protein PEX 5 to target peroxisomes for selective autophagy (pexophagy), providing a mechanism for regulation of peroxisomal homeostasis using ROS as a rheostat.
Topics: Animals; Autophagy; Homeostasis; Humans; Organelles; Peroxisomes; Signal Transduction
PubMed: 26967755
DOI: 10.1016/j.ceb.2016.02.017 -
Molecular Biology of the Cell May 2022Lysosomal degradative compartments hydrolyze macromolecules to generate basic building blocks that fuel metabolic pathways in our cells. They also remove misfolded...
Lysosomal degradative compartments hydrolyze macromolecules to generate basic building blocks that fuel metabolic pathways in our cells. They also remove misfolded proteins and control size, function, and number of cytoplasmic organelles via constitutive and regulated autophagy. These catabolic processes attract interest because their defective functioning is linked to human disease and their molecular components are promising pharmacologic targets. The capacity to quantitatively assess them is highly sought-after. Here we present a tandem-fluorescent reporter consisting of a HaloTag-GFP chimera appended at the C- or at the N-terminus of select polypeptides to monitor protein and organelle delivery to the lysosomal compartment. The Halo-GFP changes color on fluorescent pulse with cell-permeable HaloTag ligands and, again, on delivery to acidic, degradative lysosomal compartments, where the fluorescent ligand-associated HaloTag is relatively stable, whereas the GFP portion is not, as testified by loss of the green fluorescence and generation of a protease-resistant, fluorescent HaloTag fragment. The Halo-GFP tandem fluorescent reporter presented in our study allows quantitative and, crucially, time-resolved analyses of protein and organelle transport to the lysosomal compartment by high resolution confocal laser scanning microscopy, antibody-free electrophoretic techniques and flow cytometry.
Topics: Green Fluorescent Proteins; Humans; Lysosomes; Metabolic Networks and Pathways; Microscopy, Confocal; Organelles; Proteins
PubMed: 35108065
DOI: 10.1091/mbc.E21-10-0526 -
Journal of Lipid Research Aug 2018The endoplasmic reticulum (ER) is the main center for the synthesis of various lipid types in cells, and newly synthesized lipids are delivered from the ER to other... (Review)
Review
The endoplasmic reticulum (ER) is the main center for the synthesis of various lipid types in cells, and newly synthesized lipids are delivered from the ER to other organelles. In the past decade, various lipid transfer proteins (LTPs) have been recognized as mediators of lipid transport from the ER to other organelles; inter-organelle transport occurs at membrane contact sites (MCSs) and in a nonvesicular manner. Although the intermembrane transfer reaction catalyzed by LTPs is an equilibrium reaction, various types of newly synthesized lipids are transported unidirectionally in cells. This review provides a brief history of the inter-organelle trafficking of lipids and summarizes the structural and biochemical characteristics of the ceramide transport protein (CERT) as a typical LTP acting at MCSs. In addition, this review compares several LTP-mediated inter-organelle lipid trafficking systems and proposes that LTPs generate unidirectional fluxes of specific lipids between different organelles by indirect coupling with the metabolic reactions that occur in specific organelles. Moreover, the available data also suggest that the major advantage of LTP-mediated lipid transport at MCSs may be the accuracy of delivery. Finally, how cholesterol is enriched in the plasma membrane is discussed from a thermodynamic perspective.
Topics: Animals; Biological Transport; Carrier Proteins; Cell Membrane; Humans; Organelles
PubMed: 29884707
DOI: 10.1194/jlr.R085324 -
International Journal of Nanomedicine 2020Nanomedicines (NMs) have played an increasing role in cancer therapy as carriers to efficiently deliver therapeutics into tumor cells. For this application, the uptake... (Review)
Review
Nanomedicines (NMs) have played an increasing role in cancer therapy as carriers to efficiently deliver therapeutics into tumor cells. For this application, the uptake of NMs by tumor cells is usually a prerequisite to deliver the cargo to intracellular locations, which mainly relies on endocytosis. NMs can enter cells through a variety of endocytosis pathways. Different endocytosis pathways exhibit different intracellular trafficking routes and diverse subcellular localizations. Therefore, a comprehensive understanding of endocytosis mechanisms is necessary for increasing cellular entry efficiency and to trace the fate of NMs after internalization. This review focuses on endocytosis pathways of NMs in tumor cells, mainly including clathrin- and caveolae-mediated endocytosis pathways, involving effector molecules, expression difference of those molecules between normal and tumor cells, as well as the intracellular trafficking route of corresponding endocytosis vesicles. Then, the latest strategies for NMs to actively employ endocytosis are described, including improving tumor cellular uptake of NMs by receptor-mediated endocytosis, transporter-mediated endocytosis and enabling drug activity by changing intracellular routes. Finally, active targeting strategies towards intracellular organelles are also mentioned. This review will be helpful not only in explicating endocytosis and the trafficking process of NMs and elucidating anti-tumor mechanisms inside the cell but also in rendering new ideas for the design of highly efficacious and cancer-targeted NMs.
Topics: Animals; Caveolae; Clathrin; Endocytosis; Humans; Nanomedicine; Neoplasms; Organelles
PubMed: 33268987
DOI: 10.2147/IJN.S274289