-
Philosophical Transactions of the Royal... Feb 2019Macropinocytosis-the large-scale, non-specific uptake of fluid by cells-is used by Dictyostelium discoideum amoebae to obtain nutrients. These cells form circular... (Review)
Review
Macropinocytosis-the large-scale, non-specific uptake of fluid by cells-is used by Dictyostelium discoideum amoebae to obtain nutrients. These cells form circular ruffles around regions of membrane defined by a patch of phosphatidylinositol (3,4,5)-trisphosphate (PIP3) and the activated forms of the small G-proteins Ras and Rac. When this ruffle closes, a vesicle of the medium is delivered to the cell interior for further processing. It is accepted that PIP3 is required for efficient macropinocytosis. Here, we assess the roles of Ras and Rac in Dictyostelium macropinocytosis. Gain-of-function experiments show that macropinocytosis is stimulated by persistent Ras activation and genetic analysis suggests that RasG and RasS are the key Ras proteins involved. Among the activating guanine exchange factors (GEFs), GefF is implicated in macropinocytosis by an insertional mutant. The individual roles of Rho family proteins are little understood but activation of at least some may be independent of PIP3. This article is part of the Theo Murphy meeting issue 'Macropinocytosis'.
Topics: Dictyostelium; Monomeric GTP-Binding Proteins; Pinocytosis
PubMed: 30967009
DOI: 10.1098/rstb.2018.0150 -
Autophagy May 2017Macroautophagy/autophagy is vital for cellular homeostasis and helps cells respond to various stress situations. Macropinocytosis enables cells to nonselectively engulf... (Review)
Review
Macroautophagy/autophagy is vital for cellular homeostasis and helps cells respond to various stress situations. Macropinocytosis enables cells to nonselectively engulf and take up large volumes of fluid and is known to supply amino acids to cells. The stem cell-enriched limbal epithelium has the machinery necessary to carry out both autophagy and macropinocytosis; however, both processes are relatively understudied in this tissue. We have demonstrated that these processes are linked via MIR103-MIR107, a microRNA family that is limbal epithelial-preferred. Loss of MIR103-MIR107 causes the accumulation of large vacuoles that originate, in part, from a dysregulation in macropinocytosis via activation of SRC-RAS signaling. We found that these vacuoles were autophagic in nature and retained in cells due to inappropriate regulation of end-stage autophagy. Specifically, MIR103-MIR107 regulates diacylglycerol-PRKC/protein kinase C and CDK5 (cyclin dependent kinase 5) signaling, which enables DNM1 (dynamin 1) to function in vacuole clearance.
Topics: Animals; Autophagy; Epithelial Cells; Humans; MicroRNAs; Pinocytosis; Stem Cells; Vacuoles
PubMed: 28402214
DOI: 10.1080/15548627.2017.1287658 -
The FEBS Journal Nov 2017Macropinocytosis is a mechanism for the nonspecific bulk uptake and internalisation of extracellular fluid. This plays specific and distinct roles in diverse cell types... (Review)
Review
Macropinocytosis is a mechanism for the nonspecific bulk uptake and internalisation of extracellular fluid. This plays specific and distinct roles in diverse cell types such as macrophages, dendritic cells and neurons, by allowing cells to sample their environment, extract extracellular nutrients and regulate plasma membrane turnover. Macropinocytosis has recently been implicated in several diseases including cancer, neurodegenerative diseases and atherosclerosis. Uptake by macropinocytosis is also exploited by several intracellular pathogens to gain entry into host cells. Both capturing and subsequently processing large volumes of extracellular fluid poses a number of unique challenges for the cell. Macropinosome formation requires coordinated three-dimensional manipulation of the cytoskeleton to form shaped protrusions able to entrap extracellular fluid. The following maturation of these large vesicles then involves a complex series of membrane rearrangements to shrink and concentrate their contents, while delivering components required for digestion and recycling. Recognition of the diverse importance of macropinocytosis in physiology and disease has prompted a number of recent studies. In this article, we summarise advances in our understanding of both macropinosome formation and maturation, and seek to highlight the important unanswered questions.
Topics: Animals; Carrier Proteins; Cell Surface Extensions; Endosomes; Humans; Pinocytosis
PubMed: 28544479
DOI: 10.1111/febs.14115 -
Methods (San Diego, Calif.) Aug 2017Macropinocytosis is the uptake of extracellular fluid within vesicles of varying size that takes place during numerous cellular processes in a large variety of cells. A... (Review)
Review
Macropinocytosis is the uptake of extracellular fluid within vesicles of varying size that takes place during numerous cellular processes in a large variety of cells. A growing number of pathogens, including viruses, parasites, and bacteria are known to induce macropinocytosis during their entry into targeted host cells. We have recently discovered that the human enteroinvasive, bacterial pathogen Shigella causes in situ macropinosome formation during its entry into epithelial cells. These infection-associated macropinosomes are not generated to ingest the bacteria, but are instead involved in Shigella's intracellular niche formation. They make contacts with the phagocytosed shigellae to promote vacuolar membrane rupture and their cytosolic release. Here, we provide an overview of the different imaging approaches that are currently used to analyze macropinocytosis during infectious processes with a focus on Shigella entry. We detail the advantages and disadvantages of genetically encoded reporters as well as chemical probes to trace fluid phase uptake. In addition, we report how such reporters can be combined with ultrastructural approaches for correlative light electron microscopy either in thin sections or within large volumes. The combined imaging techniques introduced here provide a detailed characterization of macropinosomes during bacterial entry, which, apart from Shigella, are relevant for numerous other ones, including Salmonella, Brucella or Mycobacteria.
Topics: Bacteriological Techniques; Biomarkers; Dysentery, Bacillary; Endosomes; Host-Pathogen Interactions; Humans; Microscopy, Electron; Pinocytosis; Shigella
PubMed: 28522322
DOI: 10.1016/j.ymeth.2017.05.007 -
The Journal of Cell Biology Jul 2021Actin organization underpins conserved functions at the leading edge of cells. In this issue, Yang et al. (2021. J. Cell Biol.https://doi.org/10.1083/jcb.202010096)...
Actin organization underpins conserved functions at the leading edge of cells. In this issue, Yang et al. (2021. J. Cell Biol.https://doi.org/10.1083/jcb.202010096) characterize Leep1 as a bi-functional regulator of migration and macropinocytosis through PIP3 and the Scar/WAVE complex.
Topics: Actins; Pinocytosis
PubMed: 34128957
DOI: 10.1083/jcb.202105141 -
Advanced Drug Delivery Reviews Sep 2022Nanoparticles (NP) are attractive options for the therapeutic delivery of active pharmaceutical drugs, proteins and nucleic acids into cells, tissues and organs.... (Review)
Review
Nanoparticles (NP) are attractive options for the therapeutic delivery of active pharmaceutical drugs, proteins and nucleic acids into cells, tissues and organs. Research into the development and application of NP most often starts with a diverse group of scientists, including chemists, bioengineers and material and pharmaceutical scientists, who design, fabricate and characterize NP in vitro (Stage 1). The next step (Stage 2) generally investigates cell toxicity as well as the processes by which NP bind, are internalized and deliver their cargo to appropriate model tissue culture cells. Subsequently, in Stage 3, selected NP are tested in animal systems, mostly mouse. Whereas the chemistry-based development and analysis in Stage 1 is increasingly sophisticated, the investigations in Stage 2 are not what could be regarded as 'state-of-the-art' for the cell biology field and the quality of research into NP interactions with cells is often sub-standard. In this review we describe our current understanding of the mechanisms by which particles gain entry into mammalian cells via endocytosis. We summarize the most important areas for concern, highlight some of the most common mis-conceptions, and identify areas where NP scientists could engage with trained cell biologists. Our survey of the different mechanisms of uptake into cells makes us suspect that claims for roles for caveolae, as well as macropinocytosis, in NP uptake into cells have been exaggerated, whereas phagocytosis has been under-appreciated.
Topics: Animals; Clathrin; Endocytosis; Mammals; Mice; Nanoparticles; Pharmaceutical Preparations; Pinocytosis
PubMed: 35777667
DOI: 10.1016/j.addr.2022.114403 -
Frontiers in Immunology 2023Invariant chain (Ii, CD74) is a type II transmembrane glycoprotein that acts as a chaperone and facilitates the folding and transport of MHC II chains. By assisting the...
Invariant chain (Ii, CD74) is a type II transmembrane glycoprotein that acts as a chaperone and facilitates the folding and transport of MHC II chains. By assisting the assembly and subcellular targeting of MHC II complexes, Ii has a wide impact on the functions of antigen-presenting cells such as antigen processing, endocytic maturation, signal transduction, cell migration, and macropinocytosis. Ii is a multifunctional molecule that can alter endocytic traffic and has several interacting molecules. To understand more about Ii's function and to identify further Ii interactors, a yeast two-hybrid screening was performed. Retinoic Acid-Induced 14 (Rai14) was detected as a putative interaction partner, and the interaction was confirmed by co-immunoprecipitation. Rai14 is a poorly characterized protein, which is believed to have a role in actin cytoskeleton and membrane remodeling. In line with this, we found that Rai14 localizes to membrane ruffles, where it forms macropinosomes. Depletion of Rai14 in antigen-presenting cells delays MHC II internalization, affecting macropinocytic activity. Intriguingly, we demonstrated that, similar to Ii, Rai14 is a positive regulator of macropinocytosis and a negative regulator of cell migration, two antagonistic processes in antigen-presenting cells. This antagonism is known to depend on the interaction between myosin II and Ii. Here, we show that Rai14 also binds to myosin II, suggesting that Ii, myosin II, and Rai14 work together to coordinate macropinocytosis and cell motility.
Topics: Tretinoin; Histocompatibility Antigens Class II; Pinocytosis; Cytoskeletal Proteins; Myosin Type II
PubMed: 37545539
DOI: 10.3389/fimmu.2023.1182180 -
Philosophical Transactions of the Royal... Feb 2019In tumour cells, macropinocytosis functions as an amino acid supply route and supports cancer cell survival and proliferation. Initially demonstrated in oncogenic... (Review)
Review
In tumour cells, macropinocytosis functions as an amino acid supply route and supports cancer cell survival and proliferation. Initially demonstrated in oncogenic KRAS-driven models of pancreatic cancer, macropinocytosis triggers the internalization of extracellular proteins via discrete endocytic vesicles called macropinosomes. The incoming protein cargo is targeted for lysosome-dependent degradation, causing the intracellular release of amino acids. These protein-derived amino acids support metabolic fitness by contributing to the intracellular amino acid pools, as well as to the biosynthesis of central carbon metabolites. In this way, macropinocytosis represents a novel amino acid supply route that tumour cells use to survive the nutrient-poor conditions of the tumour microenvironment. Macropinocytosis has also emerged as an entry mechanism for a variety of nanomedicines, suggesting that macropinocytosis regulation in the tumour setting can be harnessed for the delivery of anti-cancer therapeutics. A slew of recent studies point to the possibility that macropinocytosis is a pervasive feature of many different tumour types. In this review, we focus on the role of this important uptake mechanism in a variety of cancers and highlight the main molecular drivers of macropinocytosis in these malignancies. This article is part of the Theo Murphy meeting issue 'Macropinocytosis'.
Topics: Humans; Neoplasms; Pinocytosis; Tumor Microenvironment
PubMed: 30967003
DOI: 10.1098/rstb.2018.0153 -
Sub-cellular Biochemistry 2022The distinct movements of macropinosome formation and maturation have corresponding biochemical activities which occur in a defined sequence of stages and transitions...
The distinct movements of macropinosome formation and maturation have corresponding biochemical activities which occur in a defined sequence of stages and transitions between those stages. Each stage in the process is regulated by variously phosphorylated derivatives of phosphatidylinositol (PtdIns) which reside in the cytoplasmic face of the membrane lipid bilayer. PtdIns derivatives phosphorylated at the 3' position of the inositol moiety, called 3' phosphoinositides (3'PIs), regulate different stages of the sequence. 3'PIs are synthesized by numerous phosphoinositide 3'-kinases (PI3K) and other lipid kinases and phosphatases, which are themselves regulated by small GTPases of the Ras superfamily. The combined actions of these enzymes localize four principal species of 3'PI to distinct domains of the plasma membrane or to discrete organelles, with distinct biochemical activities confined to those domains. Phosphatidylinositol (3,4,5)-trisphosphate (PtdIns(3,4,5)P) and phosphatidylinositol (3,4)-bisphosphate (PtdIns(3,4)P) regulate the early stages of macropinosome formation, which include cell surface ruffling and constrictions of circular ruffles which close into macropinosomes. Phosphatidylinositol 3-phosphate (PtdIns3P) regulates macropinosome fusion with other macropinosomes and early endocytic organelles. Phosphatidylinositol (3,5)-bisphosphate (PtdIns(3,5)P) mediates macropinosome maturation and shrinkage, through loss of ions and water, and subsequent traffic to lysosomes. The different characteristic rates of macropinocytosis in different cell types indicate levels of regulation which may be governed by the cell's capacity to generate 3'PIs.
Topics: Cell Membrane; Endosomes; Phosphatidylinositols; Phosphoric Monoester Hydrolases; Pinocytosis
PubMed: 35378706
DOI: 10.1007/978-3-030-94004-1_7 -
Arteriosclerosis, Thrombosis, and... Sep 2020Macrophages are immune cells, capable to remodel the extracellular matrix, which can harbor extracellular DNA incorporated into neutrophil extracellular traps (NETs). To... (Comparative Study)
Comparative Study
OBJECTIVE
Macrophages are immune cells, capable to remodel the extracellular matrix, which can harbor extracellular DNA incorporated into neutrophil extracellular traps (NETs). To study the breakdown of NETs we studied the capability of macrophage subsets to degrade these structures in vitro and in vivo in a murine thrombosis model. Furthermore, we analyzed human abdominal aortic aneurysm samples in support of our in vitro and in vivo results. Approach and Results: Macrophages were seeded onto blood clots or isolated NETs and polarized. All macrophages were capable to degrade NETs. For initial breakdown, macrophages relied on extracellular deoxyribonucleases. Proinflammatory polarization enhanced NET degradation. The boost in degradation was because of increased macropinocytosis, as inhibition by imipramine diminished their NET breakdown. Inhibition of macropinocytosis in a murine thrombosis model led to increased NET burden and reduced thrombus resolution in vivo. When analyzing abdominal aortic aneurysm samples, macrophage density furthermore corresponded negatively with the amount of local NETs in the intraluminal thrombi as well as in the vessel wall, as increased macrophage density was associated with a reduction in NET burden.
CONCLUSIONS
We provide evidence that macrophages degrade NETs by extracellular predigestion and subsequent uptake. Furthermore, we show that proinflammatory macrophages increase NET degradation through enhanced macropinocytosis, priming them for NET engulfment. Based on our findings, that inhibition of macropinocytosis in mice corresponded to increased NET amounts in thrombi and that local macrophage density in human abdominal aortic aneurysm is negatively associated with surrounding NETs, we hypothesize, that macrophages are able to degrade NETs in vivo.
Topics: Animals; Aortic Aneurysm, Abdominal; Cells, Cultured; Deoxyribonuclease I; Deoxyribonucleases; Disease Models, Animal; Endodeoxyribonucleases; Exodeoxyribonucleases; Extracellular Traps; Female; Humans; Imipramine; Interferon-gamma; Interleukin-13; Interleukin-4; Kinetics; Lipopolysaccharides; Macrophage Activation; Macrophages; Mice; Mice, Inbred C57BL; Muscle Proteins; Neutrophils; Phagocytosis; Phenotype; Phosphoproteins; Pinocytosis; Vena Cava, Inferior; Venous Thrombosis
PubMed: 32673525
DOI: 10.1161/ATVBAHA.120.314883