The discovery of genetic forms of Parkinson’s disease (PD) has highlighted the importance of the autophagy/lysosomal and mitochondrial/oxidative stress pathways in disease pathogenesis. However, ...recently identified PD-linked genes, including DNAJC6 (auxilin), SYNJ1 (synaptojanin 1), and the PD risk gene SH3GL2 (endophilin A1), have also highlighted disruptions in synaptic vesicle endocytosis (SVE) as a significant contributor to disease pathogenesis. Additionally, the roles of other PD genes such as LRRK2, PRKN, and VPS35 in the regulation of SVE are beginning to emerge. Here we discuss the recent work on the contribution of dysfunctional SVE to midbrain dopaminergic neurons’ selective vulnerability and highlight pathways that demonstrate the interplay of synaptic, mitochondrial, and lysosomal dysfunction in the pathogenesis of PD.
Emerging genetic and mechanistic studies link synaptic, mitochondrial, and lysosomal dysfunction as major contributors to the degeneration of midbrain dopaminergic neurons.
Recently, endocytic genes (DNAJC6, SYNJ1, and SH3GL2) have been linked to Parkinson’s disease pathogenesis or identified as a risk factor for the disease, implicating a role for impaired synaptic vesicle endocytosis (SVE) in neurodegeneration.
Among the modifiers of proteins involved in SVE are LRRK2 and parkin. LRRK2’s and parkin’s involvement in this context is through their phosphorylation and ubiquitination actions, respectively.
Dysfunction in SVE in dopaminergic neurons can lead to increased levels of unpackaged, cytosolic DA that is subject to oxidation and pathogenic downstream effects.
A potentially important aspect in the regulation of tumour metastasis is endocytosis. This process consists of internalisation of cell-surface receptors via pinocytosis, phagocytosis or ...receptor-mediated endocytosis, the latter of which includes clathrin-, caveolae- and non-clathrin or caveolae-mediated mechanisms. Endocytosis then progresses through several intracellular compartments for sorting and routing of cargo, ending in lysosomal degradation, recycling back to the cell surface or secretion. Multiple endocytic proteins are dysregulated in cancer and regulate tumour metastasis, particularly migration and invasion. Importantly, four metastasis suppressor genes function in part by regulating endocytosis, namely, the NME, KAI, MTSS1 and KISS1 pathways. Data on metastasis suppressors identify a new point of dysregulation operative in tumour metastasis, alterations in signalling through endocytosis. This review will focus on the multicomponent process of endocytosis affecting different steps of metastasis and how metastatic-suppressor genes use endocytosis to suppress metastasis.
Force generation by actin assembly shapes cellular membranes. An experimentally constrained multiscale model shows that a minimal branched actin network is sufficient to internalize endocytic pits ...against membrane tension. Around 200 activated Arp2/3 complexes are required for robust internalization. A newly developed molecule-counting method determined that ~200 Arp2/3 complexes assemble at sites of clathrin-mediated endocytosis in human cells. Simulations predict that actin self-organizes into a radial branched array with growing ends oriented toward the base of the pit. Long actin filaments bend between attachment sites in the coat and the base of the pit. Elastic energy stored in bent filaments, whose presence was confirmed by cryo-electron tomography, contributes to endocytic internalization. Elevated membrane tension directs more growing filaments toward the base of the pit, increasing actin nucleation and bending for increased force production. Thus, spatially constrained actin filament assembly utilizes an adaptive mechanism enabling endocytosis under varying physical constraints.
Phosphatidylserine (PS), a negatively charged phospholipid present predominantly at the inner leaflet of the plasma membrane, has been widely implicated in many cellular processes including membrane ...trafficking. Along this line, PS has been demonstrated to be important for endocytosis, however, the involved mechanisms remain uncertain. By monitoring clathrin‐mediated endocytosis (CME) of single vesicles in mouse chromaffin cells using cell‐attached capacitance measurements that offer millisecond time resolution, we demonstrate in the present study that the fission‐pore duration is reduced by PS addition, indicating a stimulatory role of PS in regulating the dynamics of vesicle fission during CME. Furthermore, our results show that the PS‐mediated effect on the fission‐pore duration is Ca2+‐dependent and abolished in the absence of synaptotagmin 1 (Syt1), implying that Syt1 is necessary for the stimulatory role of PS in vesicle fission during CME. Consistently, a Syt1 mutant with a defective PS–Syt1 interaction increases the fission‐pore duration. Taken together, our study suggests that PS–Syt1 interaction may be critical in regulating fission dynamics during CME.
Phosphatidylserine (PS), a negatively charged phospholipid, may be important for endocytosis, however, the involved mechanisms remain uncertain. By monitoring clathrin‐mediated endocytosis (CME) of single vesicles in mouse chromaffin cells using cell‐attached capacitance recordings, we demonstrate that the fission‐pore duration is reduced by PS addition, indicating a stimulatory role of PS in regulating the dynamics of vesicle fission. Furthermore, the PS‐mediated effect is abolished in the absence of synaptotagmin 1 (Syt1) and a Syt1 mutant with a defective PS–Syt1 interaction increases the fission‐pore duration. Our study thus suggests an importance of the PS–Syt1 interaction in regulating fission dynamics during CME.
Flaviviruses enter host cells through the process of clathrin-mediated endocytosis, and the spectrum of host factors required for this process are incompletely understood. Here we found that ...lymphocyte antigen 6 locus E (LY6E) promotes the internalization of multiple flaviviruses, including West Nile virus, Zika virus, and dengue virus. Perhaps surprisingly, LY6E is dispensable for the internalization of the endogenous cargo transferrin, which is also dependent on clathrin-mediated endocytosis for uptake. Since viruses are substantially larger than transferrin, we reasoned that LY6E may be required for uptake of larger cargoes and tested this using transferrin-coated beads of similar size as flaviviruses. LY6E was indeed required for the internalization of transferrin-coated beads, suggesting that LY6E is selectively required for large cargo. Cell biological studies found that LY6E forms tubules upon viral infection and bead internalization, and we found that tubule formation was dependent on RNASEK, which is also required for flavivirus internalization, but not transferrin uptake. Indeed, we found that RNASEK is also required for the internalization of transferrin-coated beads, suggesting it functions upstream of LY6E. These LY6E tubules resembled microtubules, and we found that microtubule assembly was required for their formation and flavivirus uptake. Since microtubule end-binding proteins link microtubules to downstream activities, we screened the three end-binding proteins and found that EB3 promotes virus uptake and LY6E tubularization. Taken together, these results highlight a specialized pathway required for the uptake of large clathrin-dependent endocytosis cargoes, including flaviviruses.
Clathrin-mediated endocytosis (CME) regulates signaling from the plasma membrane. Analysis of clathrin-coated pit (CCP) dynamics led us to propose the existence of a rate-limiting, regulatory step(s) ...that monitor the fidelity of early stages in CCP maturation. Here we show that nascent endocytic vesicles formed in mutant cells displaying rapid, dysregulated CME are defective in early endosomal trafficking, maturation and acidification, confirming the importance of this "checkpoint." Dysregulated CME also alters EGF receptor signaling and leads to constitutive activation of the protein kinase Akt. Dynamin-1, which was thought to be neuron specific, is activated by the Akt/GSK3beta signaling cascade in non-neuronal cells to trigger rapid, dysregulated CME. Acute activation of dynamin-1 in RPE cells by inhibition of GSK3beta accelerates CME, alters CCP dynamics and, unexpectedly, increases the rate of CCP initiation. CRISPR-Cas9n-mediated knockout and reconstitution studies establish that dynamin-1 is activated by Akt/GSK3beta signaling in H1299 non-small lung cancer cells. These findings provide direct evidence for an isoform-specific role for dynamin in regulating CME and reveal a feed-forward pathway that could link signaling from cell surface receptors to the regulation of CME. Synopsis Dynamin-1, previously thought to be neuron specific, is activated by an Akt/GSK3beta signaling cascade in non-neuronal cells. Dynamin-1 activation alters the rate and regulation of clathrin-mediated endocytosis, providing a feed-forward pathway between endocytosis and signaling. An endocytic checkpoint monitors the fidelity of clathrin-coated vesicle (CCV) formation. Dysregulated clathrin-mediated endocytosis (CME) alters cell signaling and proliferation. Dynamin-1 and dynamin-2 differentially regulate early stages of CME. Endogenous dynamin-1 is activated by an Akt/GSK3beta signaling cascade, illustrating a first non-neuronal role of dynamin-1.
Receptor-mediated endocytosis provides a mechanism for the selective uptake of specific molecules thereby controlling the composition of the extracellular environment and biological processes. The ...low-density lipoprotein receptor–related protein 1 (LRP1) is a widely expressed endocytic receptor that regulates cellular events by modulating the levels of numerous extracellular molecules via rapid endocytic removal. LRP1 also participates in signalling pathways through this modulation as well as in the interaction with membrane receptors and cytoplasmic adaptor proteins. LRP1 SNPs are associated with several diseases and conditions such as migraines, aortic aneurysms, cardiopulmonary dysfunction, corneal clouding, and bone dysmorphology and mineral density. Studies using Lrp1 KO mice revealed a critical, nonredundant and tissue-specific role of LRP1 in regulating various physiological events. However, exactly how LRP1 functions to regulate so many distinct and specific processes is still not fully clear. Our recent proteomics studies have identified more than 300 secreted proteins that either directly interact with LRP1 or are modulated by LRP1 in various tissues. This review will highlight the remarkable ability of this receptor to regulate secreted molecules in a tissue-specific manner and discuss potential mechanisms underpinning such specificity. Uncovering the depth of these “hidden” specific interactions modulated by LRP1 will provide novel insights into a dynamic and complex extracellular environment that is involved in diverse biological and pathological processes.
Artemisinin and its derivatives (ARTs) are the frontline drugs against malaria, but resistance is jeopardizing their effectiveness. ART resistance is mediated by mutations in the parasite's Kelch13 ...protein, but Kelch13 function and its role in resistance remain unclear. In this study, we identified proteins located at a Kelch13-defined compartment. Inactivation of eight of these proteins, including Kelch13, rendered parasites resistant to ART, revealing a pathway critical for resistance. Functional analysis showed that these proteins are required for endocytosis of hemoglobin from the host cell. Parasites with inactivated Kelch13 or a resistance-conferring Kelch13 mutation displayed reduced hemoglobin endocytosis. ARTs are activated by degradation products of hemoglobin. Hence, reduced activity of Kelch13 and its interactors diminishes hemoglobin endocytosis and thereby ART activation, resulting in parasite resistance.
Insulin signaling controls cell growth and metabolic homeostasis. Dysregulation of this pathway causes metabolic diseases such as diabetes. Insulin signaling pathways have been extensively studied. ...Upon insulin binding, the insulin receptor (IR) triggers downstream signaling cascades. The active IR is then internalized by clathrin-mediated endocytosis. Despite decades of studies, the mechanism and regulation of clathrin-mediated endocytosis of IR remain incompletely understood. Recent studies have revealed feedback regulation of IR endocytosis through Src homology phosphatase 2 (SHP2) and the mitogen-activated protein kinase (MAPK) pathway. Here we review the molecular mechanism of IR endocytosis and its impact on the pathophysiology of insulin resistance, and discuss the potential of SHP2 as a therapeutic target for type 2 diabetes.
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