The expression of some proteins in the autophagy pathway declines with age, which may impact neurodegeneration in diseases, including Alzheimer’s Disease. We have identified a novel non-canonical ...function of several autophagy proteins in the conjugation of LC3 to Rab5+, clathrin+ endosomes containing β-amyloid in a process of LC3-associated endocytosis (LANDO). We found that LANDO in microglia is a critical regulator of immune-mediated aggregate removal and microglial activation in a murine model of AD. Mice lacking LANDO but not canonical autophagy in the myeloid compartment or specifically in microglia have a robust increase in pro-inflammatory cytokine production in the hippocampus and increased levels of neurotoxic β-amyloid. This inflammation and β-amyloid deposition were associated with reactive microgliosis and tau hyperphosphorylation. LANDO-deficient AD mice displayed accelerated neurodegeneration, impaired neuronal signaling, and memory deficits. Our data support a protective role for LANDO in microglia in neurodegenerative pathologies resulting from β-amyloid deposition.
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•LC3-associated endocytosis (LANDO) requires Rubicon and ATG5, but not FIP200•LANDO is required for recycling of Aβ receptors including TREM2 in microglia.•LANDO confers protection against Aβ deposition and murine Alzheimer’s disease (AD)•Microglial LANDO protects against neuronal loss and memory impairment in murine AD
An LC3-associated endocytosis pathway involved in the recycling of amyloid receptors is essential for the clearance of amyloid aggregates by microglia in a model of AD
Endocytic trafficking underlies processes essential for plant growth and development, including the perception of and response to abiotic and extracellular stimuli, post‐Golgi and exocytic ...trafficking, and cytokinesis. Protein adaptors and regulatory factors of clathrin‐mediated endocytosis that contribute to the formation of endocytic clathrin‐coated vesicles are evolutionarily conserved. Yet, work of the last ten years has identified differences between the endocytic mechanisms of plants and Opisthokonts involving the endocytic adaptor TPLATE complex, the requirement of actin during CME, and the function of clathrin‐independent endocytosis in the uptake of plant‐specific plasma membrane proteins. Here, we review clathrin‐mediated and ‐independent pathways in plants and describe recent advances enabled by new proteomic and imaging methods, and conditional perturbation of endocytosis. In addition, we summarize the formation and trafficking of clathrin‐coated vesicles based on temporal and structural data garnered from high‐resolution quantitative imaging studies. Finally, new information about the cross‐talk between endocytosis and other endomembrane trafficking pathways and organelles will also be discussed.
Endocytic trafficking underlies processes essential for plant growth and development. This review (a) provides an overview of plant clathrin‐mediated and ‐independent endocytosis based on recent advances enabled by cutting‐edge proteomic and imaging methods and conditional perturbation of endocytosis, yielding high‐resolution structural and temporal details, and (b) discusses the cross‐talk between endocytosis and other endomembrane trafficking pathways and organelles.
Clathrin-independent endocytosis (CIE) allows internalization of plasma membrane proteins lacking clathrin-targeting sequences, such as the major histocompatibility complex class I protein (MHCI), ...into cells. After internalization, vesicles containing MHCI fuse with transferrin-containing endosomes generated from clathrin-dependent endocytosis. In HeLa cells, MHCI is subsequently routed to late endosomes or recycled back out to the plasma membrane (PM) in distinctive tubular carriers. Arf6 is associated with endosomal membranes carrying CIE cargo and expression of an active form of Arf6 leads to the generation of vacuolar structures that trap CIE cargo immediately after endocytosis, blocking the convergence with transferrin-containing endosomes. We isolated these trapped vacuolar structures and analyzed their protein composition by mass spectrometry. Here we identify and validate six new endogenous cargo proteins (CD44, CD55, CD98, CD147, Glut1, and ICAM1) that use CIE to enter cells. CD55 and Glut1 appear to closely parallel the trafficking of MHCI, merging with transferrin endosomes before entering the recycling tubules. In contrast, CD44, CD98, and CD147 appear to directly enter the recycling tubules and by-pass the merge with EEA1-positive, transferrin-containing endosomes. This divergent itinerary suggests that sorting may occur along this CIE pathway. Furthermore, the identification of new cargo proteins will assist others studying CIE in different cell types and tissues.
Mechanisms of endocytosis Doherty, Gary J; McMahon, Harvey T
Annual review of biochemistry,
01/2009, Letnik:
78
Journal Article
Recenzirano
Endocytic mechanisms control the lipid and protein composition of the plasma membrane, thereby regulating how cells interact with their environments. Here, we review what is known about mammalian ...endocytic mechanisms, with focus on the cellular proteins that control these events. We discuss the well-studied clathrin-mediated endocytic mechanisms and dissect endocytic pathways that proceed independently of clathrin. These clathrin-independent pathways include the CLIC/GEEC endocytic pathway, arf6-dependent endocytosis, flotillin-dependent endocytosis, macropinocytosis, circular doral ruffles, phagocytosis, and trans-endocytosis. We also critically review the role of caveolae and caveolin1 in endocytosis. We highlight the roles of lipids, membrane curvature-modulating proteins, small G proteins, actin, and dynamin in endocytic pathways. We discuss the functional relevance of distinct endocytic pathways and emphasize the importance of studying these pathways to understand human disease processes.
Parkinson’s disease (PD) is one of the most common neurodegenerative disorders, affecting 1–1.5% of the total population. While progress has been made in understanding the neurodegenerative ...mechanisms that lead to cell death in late stages of PD, mechanisms for early, causal pathogenic events are still elusive. Recent developments in PD genetics increasingly point at endolysosomal (E‐L) system dysfunction as the early pathomechanism and key pathway affected in PD. Clathrin‐mediated synaptic endocytosis, an integral part of the neuronal E‐L system, is probably the main early target as evident in auxilin, RME‐8, and synaptojanin‐1 mutations that cause PD. Autophagy, another important pathway in the E‐L system, is crucial in maintaining proteostasis and a healthy mitochondrial pool, especially in neurons considering their inability to divide and requirement to function an entire life‐time. PINK1 and Parkin mutations severely perturb autophagy of dysfunctional mitochondria (mitophagy), both in the cell body and synaptic terminals of dopaminergic neurons, leading to PD. Endolysosomal sorting and trafficking is also crucial, which is complex in multi‐compartmentalized neurons. VPS35 and VPS13C mutations noted in PD target these mechanisms. Mutations in GBA comprise the most common risk factor for PD and initiate pathology by compromising lysosomal function. This is also the case for ATP13A2 mutations. Interestingly, α‐synuclein and LRRK2, key proteins involved in PD, function in different steps of the E‐L pathway and target their components to induce disease pathogenesis. In this review, we discuss these E‐L system genes that are linked to PD and how their dysfunction results in PD pathogenesis.
This article is part of the Special Issue “Synuclein”.
The endolysosomal (E‐L) system is increasingly recognized as the key pathway affected in Parkinson’s disease (PD). Clathrin‐mediated synaptic endocytosis, an integral part of the neuronal E‐L system, is probably the main early target. Autophagy, another important E‐L pathway that aids in maintaining proteostasis and a healthy mitochondrial pool is affected. Other crucial pathways such as E‐L sorting and trafficking, along with lysosomal degradation are also hampered. Key PD proteins such as α‐synuclein and LRRK2 target the E‐L components to induce pathology. Here, we comprehensively discuss these mechanisms to better understand the role of E‐L system dysfunction in PD pathogenesis.
This article is part of the Special Issue “Synuclein”.
Specific antibodies that possess a subnanomolar affinity are very difficult to obtain from human naAmacrve immunoglobulin repertoires without the use of lengthy affinity optimization procedures. ...Here, we designed a hierarchical phage-displayed antibody library system to generate an enormous diversity of combinatorial Fab fragments (6A-1017) and attempted to isolate high-affinity Fabs against the human epidermal growth factor receptor (EGFR). A primary antibody library, designated HuDVFab-8L, comprising 4.5A-109 human naAmacrve heavy chains and eight unspecified human naAmacrve light chains was selected against the EGFR-Fc protein by biopanning, and four anti-EGFR Fab clones were isolated. Because one of the Fab clones, denoted EG-L2-11, recognized a native EGFR expressed on A431 cells, the heavy chain of the Fab was shuffled with a human naAmacrve light chain repertoire with a diversity of 1.4A-108 and selected a second time against the EGFR-Fc protein again. One EG-L2-11 variant, denoted EG-19-11, recognized an EGFR epitope that was almost the same as that bound by cetuximab and had a K D of approximately 540pM for soluble EGFR, which is about 7-fold higher than that of the FabC225 derived from cetuximab. This variant was also internalized by A431 cells, likely via receptor-mediated endocytosis, and it efficiently inhibited EGF-mediated tyrosine phosphorylation of the EGFR. These results demonstrate that the use of our hierarchical antibody library system is advantageous in generating fully human antibodies especially with a therapeutic purpose.
Ultrafast endocytosis generates vesicles from the plasma membrane as quickly as 50 ms in hippocampal neurons following synaptic vesicle fusion. The molecular mechanism underlying the rapid maturation ...of these endocytic pits is not known. Here we demonstrate that synaptojanin-1, and its partner endophilin-A, function in ultrafast endocytosis. In the absence of synaptojanin or endophilin, the membrane is rapidly invaginated, but pits do not become constricted at the base. The 5-phosphatase activity of synaptojanin is involved in formation of the neck, but 4-phosphatase is not required. Nevertheless, these pits are eventually cleaved into vesicles; within a 30-s interval, synaptic endosomes form and are resolved by clathrin-mediated budding. Then synaptojanin and endophilin function at a second step to aid with the removal of clathrin coats from the regenerated vesicles. These data together suggest that synaptojanin and endophilin can mediate membrane remodeling on a millisecond timescale during ultrafast endocytosis.
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•Synaptojanin and endophilin accelerate ultrafast endocytosis•Neck formation requires endophilin and the 5-phosphatase activity of synaptojanin•Synaptic vesicles still form from synaptic endosomes in absence of these proteins•Endophilin and 4- and 5-phosphatase are required for clathrin uncoating
Ultrafast endocytosis is a distinct form of synaptic vesicle recovery occurring within milliseconds of vesicle fusion. Using flash-and-freeze electron microscopy, Watanabe et al. demonstrate dual roles for synaptojanin and endophilin in membrane remodeling and clathrin uncoating on rapid timescales during ultrafast endocytosis.
Synaptic vesicle recycling is essential for maintaining normal synaptic function. The coupling of exocytosis and endocytosis is assumed to be Ca2+ dependent, but the exact role of Ca2+ and its key ...effector synaptotagmin-1 (syt1) in regulation of endocytosis is poorly understood. Here, we probed the role of syt1 in single- as well as multi-vesicle endocytic events using high-resolution optical recordings. Our experiments showed that the slowed endocytosis phenotype previously reported after syt1 loss of function can also be triggered by other manipulations that promote asynchronous release such as Sr2+ substitution and complexin loss of function. The link between asynchronous release and slowed endocytosis was due to selective targeting of fused synaptic vesicles toward slow retrieval by the asynchronous release Ca2+ sensor synaptotagmin-7. In contrast, after single synaptic vesicle fusion, syt1 acted as an essential determinant of synaptic vesicle endocytosis time course by delaying the kinetics of vesicle retrieval in response to increasing Ca2+ levels.
•Loss of syt1, complexins, or Sr2+ substitution slows endocytosis after activity•Asynchronously released synaptic vesicles are retrieved via slow endocytosis•Link between asynchronous release and slowed endocytosis requires synaptotagmin-7•Whereas, after synchronous single synaptic vesicle fusion, syt1 delays endocytosis
In this study, Li and colleagues demonstrate that synaptic vesicle fusion machinery, comprised of synaptotagmin-1, complexins, and synaptotagmin-7, in addition to determining the timing and Ca2+ dependence of neurotransmitter release, also dictates the properties of subsequent synaptic vesicle endocytosis.
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