In cell biology, it is a vesicle formed around a particle enveloped by a phagocyte through phagocytosis.
A phagosome is formed by the fusion of the cell membrane around a microorganism, a senescent cell, or an apoptotic cell .
Phagosomes have membrane-bound proteins to recruit and fuse with lysosomes to form mature phagolysosomes.
Lysosomes contain hydrolytic enzymes and reactive oxygen species (SRO) that kill and digest pathogens.
Phagosomes can also form into non-professional phagocytes, but they can only encompass a smaller range of particles and do not contain reactive oxygen species.
Useful materials (eg, amino acids) from the digested particles move into the cytosol, and waste is removed by exocytosis .
Phagosome formation is crucial for tissue homeostasis and both innate and adaptive host defense against pathogens.
However, some bacteria can exploit phagocytosis as an invasion strategy.
They either reproduce within the phagolysosome (eg, Coxiella spp.) Or escape into the cytoplasm before the phagosomes fuse with the lysosome (eg, Rickettsia spp.).
Many mycobacteria, including Mycobacterium tuberculosis and Mycobacterium avium paratuberculosis, can manipulate the host macrophage to prevent lysosomes from fusing with phagosomes and creating mature phagolysosomes.
Such incomplete maturation of the phagosome maintains a favorable environment for pathogens within it.
Phagosomes are large enough to degrade entire bacteria, or apoptotic and senescent cells, which are generally> 0.5μm in diameter.
This means that a phagosome is several orders of magnitude larger than an endosome, which is measured in nanometers.
Phagosomes are formed when pathogens or opsonins bind to a transmembrane receptor, which are randomly distributed on the surface of phagocyte cells.
After attachment, “outside-in” signaling triggers actin polymerization and pseudopod formation, which surrounds and fuses behind the organism.
Protein kinase C, phosphoinositide 3-kinase, and phospholipase C (PLC) are all required to signal and control particle internalization.
More cell surface receptors can bind to the particle in a zipper-like mechanism as the pathogen is surrounded, increasing the avidity of attachment.
The Fc receptor (FcR), complement receptors (RC), mannose receptor, and Dectin-1 are phagocytic receptors, meaning that they can induce phagocytosis if they are expressed in non-phagocytic cells such as fibroblasts.
Other proteins, such as Toll-like receptors, are involved in pathogen pattern recognition and are often recruited to phagosomes, but do not specifically trigger phagocytosis in non-phagocytic cells, so they are not considered phagocytic receptors .
Opsonins are molecular tags, such as antibodies and complements, that bind to pathogens and upregulate phagocytosis. Immunoglobulin G (IgG) is the main type of antibody present in serum.
It is part of the adaptive immune system, but it is linked to the innate response by recruiting macrophages to engulf pathogens. The antibody binds to microbes with the Fab variable domain, and the Fc domain binds to Fc receptors (FcRs) to induce phagocytosis.
Complement-mediated internalization has much less significant membrane overhangs, but downstream signaling from both pathways converge to activate Rho GTPases.
They control the actin polymerization that is required for the phagosome to fuse with endosomes and lysosomes.
Other non-professional phagocytes have some degree of phagocytic activity, such as thyroid and bladder epithelial cells that can encompass erythrocytes and retinal epithelial cells that internalize the retinal rods.
However, non-professional phagocytes do not express specific phagocytic receptors, such as the Fc receptor, and have a much lower rate of internalization.
Some invasive bacteria can also induce phagocytosis in non-phagocytic cells to mediate host uptake. For example, Shigella can secrete toxins that disrupt the host’s cytoskeleton and enter the basolateral side of enterocytes.
Since the phagosome membrane is formed by fusion of the plasma membrane, the basic composition of the phospholipid bilayer is the same.
The endosomes and lysosomes then fuse with the phagosome to contribute to the membrane, especially when the enveloped particle is very large, such as a parasite.
They also deliver various membrane proteins to the phagosome and modify the structure of the organelle.
Phagosomes can engulf low-density latex artificial beads and then be purified along a sucrose concentration gradient, allowing structure and composition to be studied.
By purifying the phagosomes at different time points, the maturation process can also be characterized. The first phagosomes are characterized by Rab5, which transitions to Rab7 as the vesicle matures into late phagosomes.
The nascent phagosome is not inherently bactericidal. As it matures, it becomes more acidic from pH 6.5 to pH 4, gaining characteristic protein markers and hydrolytic enzymes.
Different enzymes work at various optimal pHs, forming a range for each to work in narrow stages of the ripening process. The activity of the enzyme can be adjusted by changing the pH level, allowing for greater flexibility.
The phagosome moves along the microtubules of the cytoskeleton, fusing with endosomes and lysosomes sequentially in a dynamic “kiss and flight” manner.
This intracellular transport depends on the size of the phagosomes.
Larger organelles (approximately 3 μm in diameter) are transported very persistently from the periphery of the cell to the perinuclear region.
While the smaller organelles (with a diameter of approximately 1 μm) are transported more bidirectionally between the cell center and the cell periphery.
Vacuolar proton pumps (v-ATPase) are delivered to the phagosome to acidify the organelle compartment, creating a more hostile environment for pathogens and facilitating protein degradation.
Bacterial proteins denature at low pH and become more accessible to proteases, which are unaffected by the acidic environment.
The enzymes are later recycled from the phagolysosome before egestion so that they are not wasted. The composition of the phospholipid membrane also changes as the phagosome matures.
Fusion can take from minutes to hours depending on the content of the phagosome; FcR or mannose receptor-mediated fusion takes less than 30 minutes, but phagosomes containing latex beads can take several hours to fuse with lysosomes.
It is suggested that the composition of the phagosome membrane affects the rate of maturation.
Mycobacterium tuberculosis has a highly hydrophobic cell wall, which is hypothesized to prevent membrane recycling and recruitment of fusion factors, so the phagosome does not fuse with the lysosomes and the bacterium prevents degradation.
Smaller lumenal molecules are transferred by fusion faster than larger molecules, suggesting that a small aqueous channel forms between the phagosome and other vesicles during the ‘kiss and flight’, through which only the exchange is allowed limited.
Shortly after internalization, F-actin depolymerizes from the newly formed phagosome, thus making endosomes accessible for protein fusion and delivery.
The maturation process is divided into early and late stages depending on the characteristic protein markers, regulated by small Rab GTPases.
Rab5 is present in early phagosomes, and controls the transition to late Rab7-labeled phagosomes.
Rab5 recruits PI-3 kinase and other anchor proteins such as Vps34 to the phagosome membrane, whereby endosomes can deliver proteins to the phagosome.
Rab5 partially participates in the transition to Rab7, through the CORVET complex and the HOPS complex in yeast.
The exact maturation pathway in mammals is not well understood, but it is suggested that HOPS may bind Rab7 and displace the guanosine nucleotide dissociation inhibitor (GDI). Rab11 is involved in membrane recycling.
The phagosome fuses with lysosomes to form a phagolysosome, which has various bactericidal properties.
The phagolysosome contains reactive oxygen and nitrogen species (ROS and RNS) and hydrolytic enzymes.
The compartment is also acidic due to proton pumps (v-ATPases) that transport H + across the membrane, which is used to denature bacterial proteins.
The exact properties of phagolysosomes vary depending on the type of phagocyte. Those in dendritic cells have weaker bactericidal properties than those of macrophages and neutrophils.
In addition, macrophages are divided into proinflammatory “killer” M1 and “repair” M2.
M1 phagolysosomes can metabolize arginine to highly reactive nitric oxide, while M2 uses arginine to produce ornithine to promote cell proliferation and tissue repair.
Macrophages and neutrophils are professional phagocytes in charge of most of the degradation of pathogens, but they have different bactericidal methods.
Neutrophils have granules that fuse with the phagosome. The granules contain NADPH oxidase and myeloperoxidase, which produce toxic oxygen and chlorine derivatives to kill pathogens in an oxidative burst.
Antimicrobial proteases and peptides are also released in the phagolysosome. Macrophages lack granules and rely more on acidification from phagolysosomes, glycosidases, and proteases to digest microbes.
Phagosomes in dendritic cells are less acidic and have a much weaker hydrolytic activity, due to a lower concentration of lysosomal proteases and even the presence of protease inhibitors.
Phagosome formation is linked to inflammation through common signaling molecules. PI-3 kinase and phospholipase C are involved in both the internalization mechanism and the activation of inflammation.
The two proteins, along with the Rho GTPases, are important components of the innate immune response, inducing cytokine production and activating the MAP kinase signaling cascade.
If it produces citoquinas proinflamatorias, it includes IL-1β, IL-6, TNFα and IL-12.
The process is strictly regulated and the inflammatory response varies according to the type of particle within the phagosome.
Apoptotic cells infected with pathogens will trigger inflammation, but damaged cells that degrade as part of normal tissue turnover do not. The response also differs according to opsonin-mediated phagocytosis.
FcR and mannose receptor-mediated reactions produce pro-inflammatory reactive oxygen species and arachidonic acid molecules, but complement receptor-mediated reactions do not result in these products.
Immature dendritic cells (ICD) can phagocytose, but mature dendritic cells cannot due to changes in Rho GTPases involved in remodeling of the cytoskeleton.
Dendritic cell phagosomes are less hydrolytic and acidic than those of macrophages and neutrophils, since dendritic cells are primarily involved in antigen presentation rather than pathogen degradation.
They need to retain protein fragments of a suitable size for specific bacterial recognition, so the peptides are only partially degraded.
Bacterial peptides are trafficked to the major histocompatibility complex (MHC).
Peptide antigens are presented to lymphocytes, where they bind to T cell receptors and activate T cells, bridging the gap between innate and adaptive immunity.
This is specific to mammals, birds, and jawed fish, as insects do not have adaptive immunity.
Ancient single-celled organisms like the amoeba use phagocytosis as a way to acquire nutrients, rather than an immune strategy.
They engulf other smaller microbes and digest them within the phagosome at around one bacterium per minute, which is much faster than professional phagocytes.
For the soil amoeba Dictyostelium discoideum, its main food source is the bacterium Legionella pneumophila, which causes Legionnaires’ disease in humans.
The maturation of the phagosome in amoeba is very similar to that of macrophages, so they are used as a model organism to study the process.
Phagosomes degrade senescent and apoptotic cells to maintain tissue homeostasis. Red blood cells have one of the highest turnover rates in the body, and are phagocytosed by macrophages in the liver and spleen.
In the embryo, the process of removing dead cells is not well characterized, but it is not performed by macrophages or other cells derived from hematopoietic stem cells.
It is only in the adult that apoptotic cells are phagocytosed by professional phagocytes.
Inflammation is only triggered by certain pathogen or damage associated molecular patterns (DAMP) or pathogen associated molecular patterns (PAMP), the elimination of senescent cells is not inflammatory.
Autophagosomes are different from phagosomes in that they are primarily used to selectively degrade damaged cytosolic organelles, such as mitochondria (mitophagy).
However, when the cell is starved or stressed, autophagosomes can also non-selectively degrade organelles to provide the cell with amino acids and other nutrients.
Autophagy is not limited to professional phagocytes, but was first discovered in rat hepatocytes by cell biologist Christian de Duve.
Autophagosomes have a double membrane, the inner membrane of the engulfed organelle, and it is speculated that the outer membrane will form from the endoplasmic reticulum or the ER-Golgi intermediate compartment (ERGIC).
The autophagosome also fuses with lysosomes to degrade their content. When M. tuberculosis inhibits acidification of the phagosome, gamma interferon can induce autophagy and rescue the maturation process.
Bacterial avoidance and manipulation
Many bacteria have evolved to evade the bactericidal properties of phagosomes or even exploit phagocytosis as an invasion strategy.
Mycobacterium tuberculosis targets M2 macrophages in the lower parts of the respiratory tract, which do not produce reactive oxygen species.
M. tuberculosis can also manipulate signaling pathways by secreting phosphatases such as PtpA and SapM, which disrupt protein recruitment and block phagosome acidification.
Legionella pneumophila can reshape the phagosome membrane to mimic vesicles in other parts of the secretory pathway, thus lysosomes do not recognize the phagosome and do not fuse with it.
The bacteria secrete toxins that interfere with host trafficking, so the Legionella-containing vacuole recruits membrane proteins that are generally found in the endoplasmic reticulum or ER-Golgi intermediate compartment.
This redirects the secretory vesicles to the modified phagosome and delivers nutrients to the bacteria.
Listeria monocytogenes secretes a pore-forming protein listeriolysin O so that the bacteria can escape from the phagosome into the cytosol. Listeriolysin is activated by the acidic environment of the phagosome.
In addition, Listeria secretes two phospholipase C enzymes that facilitate the escape of the phagosome.
Final comments and perspectives for future research
The dynamic and profound changes that phagosomes undergo during their maturation allow immune cells to maintain homeostasis and respond rapidly to microbial threats.
Therefore, the maturation of the phagosome has a direct impact on the outcome of immune responses and is regulated not only at the cellular level, but also at the organelle level.
In recent years, a growing body of evidence has shown various links between the phagosome maturation machinery and different signaling pathways.
Furthermore, the polarization of phagocyte cell populations allows the innate immune system to initiate and form immune responses.
For example, M1-like and M2-like phenotypes during MΦ polarization can induce different characteristics of phagosome maturation, even when found in the same tissue.
Interestingly, these characteristics are not restricted to MΦs, as is well demonstrated by the emerging concept of neutrophil polarization.
Phagosomal maturation kinetics are susceptible to various stimuli ranging from pathogen-associated cytokines and molecular patterns to damage-associated molecular patterns and opsonins.
Other factors, such as the duration of stimulation and the type of phagocyte involved, also influence phagosomal fate.
Therefore, phagosome maturation can be stimulated to enhance pathogen kill or to prevent self-peptide presentation and autoimmunity.
In contrast, phagosome maturation can be delayed to preserve pathogenic peptides for presentation to T cells to efficiently induce adaptive immunity.
More work is needed to identify the signaling pathways to and from the phagosome and to better understand how different phagocytes regulate these aspects at the molecular level.
It is currently not known whether immune signals modulate phagosome maturation by activating a single signal transduction pathway or whether multiple pathways are involved.
Furthermore, the effects of other types of environmental stimuli (eg allergens) have not yet been studied.
Based on current progress, the idea arises that phagosomes function as signaling platforms that integrate multiple intraphagosomal, intracellular and extracellular signals, which can modulate phagosome maturation.
Proteomic studies revealed that many proteins known to be involved in signaling, such as receptors and kinases, are present in phagosomes.
The phosphorylation state of these proteins is only one aspect that influences the maturation of the phagosome.
Finally, the various interactions between phagosomes, the inflammasome, and surrounding organelles, such as autophagosomes and mitochondria, suggest that the phagosome is not a solitary organelle and can communicate with other organelles.
Future research could focus on these interactions and the impact of immune signals, especially when phagosome maturation is impaired by pathogens to evade host immunity.
This could help develop new therapies that improve pathogen clearance and adaptive immunity.