The mechanistic target of rapamycin (mTOR) coordinates eukaryotic cell growth and metabolism with environmental inputs, including nutrients and growth factors. Extensive research over the past two decades has established a central role for mTOR in regulating many fundamental cell processes, from protein synthesis to autophagy, and deregulated mTOR signaling is implicated in the progression of cancer and diabetes, as well as the aging process. Here, we review recent advances in our understanding of mTOR function, regulation, and importance in mammalian physiology. We also highlight how the mTOR signaling network contributes to human disease and discuss the current and future prospects for therapeutically targeting mTOR in the clinic.

mTOR Signaling in Growth, Metabolism, and Disease

During late endosome maturation, cargo molecules are sorted into intralumenal vesicles (ILVs) of multivesicular endosomes (MVEs), and are either delivered to lysosomes for degradation or fused with the plasma membranes for exosome release. The mechanism underlying formation of exosomal ILVs and cargo sorting into ILVs destined for exosome release is still unclear. Here we show that inhibitory G protein (Gi)-coupled sphingosine 1-phosphate (S1P) receptors regulate exosomal MVE maturation. Gi-coupled S1P receptors on MVEs are constitutively activated through a constant supply of S1P via autocrine activation within organelles. We also found that the continuous activation of Gi-coupled S1P receptors on MVEs is essential for cargo sorting into ILVs destined for exosome release. Our results reveal a mechanism underlying ESCRT-independent maturation of exosomal MVEs.

Ongoing activition of sphingosine 1-phosphate receptors mediates maturation of exosomal multivesicular endosomes

Beyond the consolidated role in degrading and recycling cellular waste, the autophagic- and endo-lysosomal systems play a crucial role in extracellular release pathways. Lysosomal exocytosis is a process leading to the secretion of lysosomal content upon lysosome fusion with plasma membrane and is an important mechanism of cellular clearance, necessary to maintain cell fitness. Exosomes are a class of extracellular vesicles originating from the inward budding of the membrane of late endosomes, which may not fuse with lysosomes but be released extracellularly upon exocytosis. In addition to garbage disposal tools, they are now considered a cell-to-cell communication mechanism. Autophagy is a cellular process leading to sequestration of cytosolic cargoes for their degradation within lysosomes. However, the autophagic machinery is also involved in unconventional protein secretion and autophagy-dependent secretion, which are fundamental mechanisms for toxic protein disposal, immune signalling and pathogen surveillance. These cellular processes underline the crosstalk between the autophagic and the endosomal system and indicate an intersection between degradative and secretory functions. Further, they suggest that the molecular mechanisms underlying fusion, either with lysosomes or plasma membrane, are key determinants to maintain cell homeostasis upon stressing stimuli. When they fail, the accumulation of undigested substrates leads to pathological consequences, as indicated by the involvement of autophagic and lysosomal alteration in human diseases, namely lysosomal storage disorders, age-related neurodegenerative diseases and cancer. In this paper, we reviewed the current knowledge on the functional role of extracellular release pathways involving lysosomes and the autophagic- and endo-lysosomal systems, evaluating their implication in health and disease.

https://www.mdpi.com/1422-0067/21/7/2576/pdf

Lysosomal Exocytosis, Exosome Release and Secretory Autophagy: The Autophagic- and Endo-Lysosomal Systems Go Extracellular.

https://www.mdpi.com/2076-393X/7/3/102/pdf

The Role of Extracellular Vesicles in Viral Infection and Transmission.

Knowledge of the host response to the novel coronavirus SARS-CoV-2 remains limited, hindering the understanding of COVID-19 pathogenesis and the development of therapeutic strategies. During the course of a viral infection, host cells release exosomes and other extracellular vesicles carrying viral and host components that can modulate the immune response. The present study used a shotgun proteomic approach to map the host circulating exosomes' response to SARS-CoV-2 infection. We investigated how SARS-CoV-2 infection modulates exosome content, exosomes' involvement in disease progression, and the potential use of plasma exosomes as biomarkers of disease severity. A proteomic analysis of patient-derived exosomes identified several molecules involved in the immune response, inflammation, and activation of the coagulation and complement pathways, which are the main mechanisms of COVID-19-associated tissue damage and multiple organ dysfunctions. In addition, several potential biomarkers-such as fibrinogen, fibronectin, complement C1r subcomponent and serum amyloid P-component-were shown to have a diagnostic feature presenting an area under the curve (AUC) of almost 1. Proteins correlating with disease severity were also detected. Moreover, for the first time, we identified the presence of SARS-CoV-2 RNA in the exosomal cargo, which suggests that the virus might use the endocytosis route to spread infection. Our findings indicate circulating exosomes' significant contribution to several processes-such as inflammation, coagulation, and immunomodulation-during SARS-CoV-2 infection. The study's data are available via ProteomeXchange with the identifier PXD021144.

/pdf/circulating-exosomes-are-strongly-involved-in-sars-cov-2-18rtg4rovt.pdf

Circulating Exosomes Are Strongly Involved in SARS-CoV-2 Infection

Lysosomes are acidic cell compartments containing a large set of hydrolytic enzymes. These lysosomal hydrolases degrade proteins, lipids, polysaccharides, and nucleic acids into their constituents. Materials to be degraded can reach lysosomes either from inside the cell, by autophagy, or from outside the cell, by different forms of endocytosis. In addition to their degradative functions, lysosomes are also able to extracellularly release their contents by lysosomal exocytosis. These organelles move from the perinuclear region along microtubules towards the proximity of the plasma membrane, then the lysosomal and plasma membrane fuse together via a Ca2+-dependent process. The fusion of the lysosomal membrane with plasma membrane plays an important role in plasma membrane repair, while the secretion of lysosomal content is relevant for the remodelling of extracellular matrix and release of functional substrates. Lysosomal storage disorders (LSDs) and age-related neurodegenerative disorders, such as Parkinson's and Alzheimer's diseases, share as a pathological feature the accumulation of undigested material within organelles of the endolysosomal system. Recent studies suggest that lysosomal exocytosis stimulation may have beneficial effects on the accumulation of these unprocessed aggregates, leading to their extracellular elimination. However, many details of the molecular machinery required for lysosomal exocytosis are only beginning to be unravelled. Here, we are going to review the current literature on molecular mechanisms and biological functions underlying lysosomal exocytosis, to shed light on the potential of lysosomal exocytosis stimulation as a therapeutic approach.

https://www.mdpi.com/2077-0375/10/12/406/pdf

Lysosomal Exocytosis: The Extracellular Role of an Intracellular Organelle.

Under physio-pathological conditions, cells release membrane-surrounded structures named Extracellular Vesicles (EVs), which convey their molecular cargo to neighboring or distant cells influencing their metabolism. Besides their involvement in the intercellular communication, EVs might represent a tool used by cells to eliminate unnecessary/toxic material. Here, we revised the literature exploring the link between EVs and redox biology. The first proof of this link derives from evidence demonstrating that EVs from healthy cells protect target cells from oxidative insults through the transfer of antioxidants. Oxidative stress conditions influence the release and the molecular cargo of EVs that, in turn, modulate the redox status of target cells. Oxidative stress-related EVs exert both beneficial or harmful effects, as they can carry antioxidants or ROS-generating enzymes and oxidized molecules. As mediators of cell-to-cell communication, EVs are also implicated in the pathophysiology of oxidative stress-related diseases. The review found evidence that numerous studies speculated on the role of EVs in redox signaling and oxidative stress-related pathologies, but few of them unraveled molecular mechanisms behind this complex link. Thus, the purpose of this review is to report and discuss this evidence, highlighting that the analysis of the molecular content of oxidative stress-released EVs (reminiscent of the redox status of originating cells), is a starting point for the use of EVs as diagnostic and therapeutic tools in oxidative stress-related diseases.

Extracellular Vesicles under Oxidative Stress Conditions: Biological Properties and Physiological Roles

Extracellular vesicles (EVs) have received increasing attention over the last two decades. Initially, they were considered as just a garbage disposal tool; however, it has progressively become clear that their protein, nucleic acid (namely miRNA and mRNA), and lipid contents have signaling functions. Besides, it has been established that cells release different types of vesicular structures for which characterization is still in its infancy. Many stress conditions, such as hypoxia, senescence, and oncogene activation have been associated with the release of higher levels of EVs. Further, evidence has shown that autophagic–lysosomal pathway abnormalities also affect EV release. In fact, in neurodegenerative diseases characterized by the accumulation of toxic proteins, although it has not become clear to what extent the intracellular storage of undigested materials itself has beneficial/adverse effects, these proteins have also been shown to be released extracellularly via EVs. Lysosomal storage disorders (LSDs) are characterized by accumulation of undigested substrates within the endosomal–lysosomal system, due either to genetic mutations in lysosomal proteins or to treatment with pharmacological agents. Here, we review studies investigating the role of lysosomal and autophagic dysfunction on the release of EVs, with a focus on studies exploring the release of EVs in LSD models of both genetic and pharmacological origin. A better knowledge of EV-releasing pathways activated in lysosomal stress conditions will provide information on the role of EVs in both alleviating intracellular storage of undigested materials and spreading the pathology to the neighboring tissue.

https://www.mdpi.com/2073-4425/10/7/510/pdf

Federica Delo

Papers

Lysosomal Exocytosis, Exosome Release and Secretory Autophagy: The Autophagic- and Endo-Lysosomal Systems Go Extracellular.

The Role of Extracellular Vesicles in Viral Infection and Transmission.

Lysosomal Exocytosis: The Extracellular Role of an Intracellular Organelle.

Extracellular Vesicles under Oxidative Stress Conditions: Biological Properties and Physiological Roles

Insight into the Role of Extracellular Vesicles in Lysosomal Storage Disorders