Deacidification of endolysosomes by neuronal aging drives synapse loss.

Previously, we found that age-dependent accumulation of beta-amyloid is not sufficient to cause synaptic decline. Late-endocytic organelles (LEOs) may be driving synaptic decline as lysosomes (Lys) are a target of cellular aging and relevant for synapses. We found that LAMP1-positive LEOs increased in size and number and accumulated near synapses in aged neurons and brains. LEOs' distal accumulation might relate to the increased anterograde movement in aged neurons. Dissecting the LEOs, we found that late-endosomes accumulated while there are fewer terminal Lys in aged neurites, but not in the cell body. The most abundant LEOs were degradative Lys or endolysosomes (ELys), especially in neurites. ELys activity was reduced because of acidification defects, supported by the reduction in v-ATPase subunit V0a1 with aging. Increasing the acidification of aged ELys recovered degradation and reverted synaptic decline, while alkalinization or v-ATPase inhibition, mimicked age-dependent Lys and synapse dysfunction. We identify ELys deacidification as a neuronal mechanism of age-dependent synapse loss. Our findings suggest that future therapeutic strategies to address endolysosomal defects might be able to delay age-related synaptic decline.

Delay of EGF-Stimulated EGFR Degradation in Myotonic Dystrophy Type 1 (DM1).

Myotonic dystrophy type 1 (DM1) is an autosomal dominant disease caused by a CTG repeat expansion in the 3' untranslated region of the dystrophia myotonica protein kinase gene. AKT dephosphorylation and autophagy are associated with DM1. Autophagy has been widely studied in DM1, although the endocytic pathway has not. AKT has a critical role in endocytosis, and its phosphorylation is mediated by the activation of tyrosine kinase receptors, such as epidermal growth factor receptor (EGFR). EGF-activated EGFR triggers the internalization and degradation of ligand-receptor complexes that serve as a PI3K/AKT signaling platform. Here, we used primary fibroblasts from healthy subjects and DM1 patients. DM1-derived fibroblasts showed increased autophagy flux, with enlarged endosomes and lysosomes. Thereafter, cells were stimulated with a high concentration of EGF to promote EGFR internalization and degradation. Interestingly, EGF binding to EGFR was reduced in DM1 cells and EGFR internalization was also slowed during the early steps of endocytosis. However, EGF-activated EGFR enhanced AKT and ERK1/2 phosphorylation levels in the DM1-derived fibroblasts. Therefore, there was a delay in EGF-stimulated EGFR endocytosis in DM1 cells; this alteration might be due to the decrease in the binding of EGF to EGFR, and not to a decrease in AKT phosphorylation.

Urolithin B: Two-way attack on IAPP proteotoxicity with implications for diabetes.

Introduction: Diabetes is one of the major metabolic diseases worldwide. Despite being a complex systemic pathology, the aggregation and deposition of Islet Amyloid Polypeptide (IAPP), or amylin, is a recognized histopathological marker of the disease. Although IAPP proteotoxicity represents an important trigger of ß-cell dysfunction and ultimately death, its exploitation as a therapeutic tool remains underdeveloped. The bioactivity of (poly)phenols towards inhibition of pathological protein aggregation is well known, however, most of the identified molecules have limited bioavailability. Methods: Using a strategy combining in silico, cell-free and cell studies, we scrutinized a unique in-house collection of (poly)phenol metabolites predicted to appear in the human circulation after (poly)phenols ingestion. Results: We identified urolithin B as a potent inhibitor of IAPP aggregation and a powerful modulator of cell homeostasis pathways. Urolithin B was shown to affect IAPP aggregation pattern, delaying the formation of amyloid fibrils and altering their size and morphology. The molecular mechanisms underlying urolithin B-mediated protection include protein clearance pathways, mitochondrial function, and cell cycle ultimately rescuing IAPP-mediated cell dysfunction and death. Discussion: In brief, our study uncovered urolithin B as a novel small molecule targeting IAPP pathological aggregation with potential to be exploited as a therapeutic tool for mitigating cellular dysfunction in diabetes. Resulting from the colonic metabolism of dietary ellagic acid in the human body, urolithin B bioactivity has the potential to be explored in nutritional, nutraceutical, and pharmacological perspectives.

Weibel-Palade bodies at a glance.

The vascular environment can rapidly alter, and the speed with which responses to both physiological and pathological changes are required necessitates the existence of a highly responsive system. The endothelium can quickly deliver bioactive molecules by regulated exocytosis of its secretory granules, the Weibel-Palade bodies (WPBs). WPBs include proteins that initiate both haemostasis and inflammation, as well those that modulate blood pressure and angiogenesis. WPB formation is driven by von Willebrand factor, their most abundant protein, which controls both shape and size of WPBs. WPB are generated in a range of sizes, with the largest granules over ten times the size of the smallest. In this Cell Science at a Glance and the accompanying poster, we discuss the emerging mechanisms by which WPB size is controlled and how this affects the ability of this organelle to modulate haemostasis. We will also outline the different modes of exocytosis and their polarity that are currently being explored, and illustrate that these large secretory organelles provide a model for how elements of secretory granule biogenesis and exocytosis cooperate to support a complex and diverse set of functions.

Type II PI4-kinases control Weibel-Palade body biogenesis and von Willebrand factor structure in human endothelial cells.

Weibel-Palade bodies (WPBs) are endothelial storage organelles that mediate the release of molecules involved in thrombosis, inflammation and angiogenesis, including the pro-thrombotic glycoprotein von Willebrand factor (VWF). Although many protein components required for WPB formation and function have been identified, the role of lipids is almost unknown. We examined two key phosphatidylinositol kinases that control phosphatidylinositol 4-phosphate levels at the trans-Golgi network, the site of WPB biogenesis. RNA interference of the type II phosphatidylinositol 4-kinases PI4KIIα and PI4KIIß in primary human endothelial cells leads to formation of an increased proportion of short WPB with perturbed packing of VWF, as exemplified by increased exposure of antibody-binding sites. When stimulated with histamine, these cells release normal levels of VWF yet, under flow, form very few platelet-catching VWF strings. In PI4KIIα-deficient mice, immuno-microscopy revealed that VWF packaging is also perturbed and these mice exhibit increased blood loss after tail cut compared to controls. This is the first demonstration that lipid kinases can control the biosynthesis of VWF and the formation of WPBs that are capable of full haemostatic function.

The host endocytic pathway is essential for Plasmodium berghei late liver stage development.

The obligate intracellular liver stage of the Plasmodium parasite represents a bottleneck in the parasite life cycle and remains a promising target for therapeutic intervention. During this stage, parasites undergo dramatic morphological changes and achieve one of the fastest replication rates among eukaryotic species. Nevertheless, relatively little is known about the parasite interactions with the host hepatocyte. Using immunofluorescence, live cell imaging and electron microscopy, we show that Plasmodium berghei parasites are surrounded by vesicles from the host late endocytic pathway. We found that these vesicles are acidic and contain the membrane markers Rab7a, CD63 and LAMP1. When host cell vesicle acidification was disrupted using ammonium chloride or Concanamycin A during the late liver stage of infection, parasite survival was not affected, but schizont size was significantly decreased. Furthermore, when the host cell endocytic pathway was loaded with BSA-gold, gold particles were found within the parasite cytoplasm, showing the transport of material from the host endocytic pathway toward the parasite interior. These observations reveal a novel Plasmodium-host interaction and suggest that vesicles from the host endolysosomal pathway could represent an important source of nutrients exploited by the fast-growing late liver stage parasites.