Endocytosis

EphB6 is an understudied ephrin receptor tyrosine pseudokinase that is downregulated in multiple types of metastatic cancers. Unlike its kinase-active counterparts which autophosphorylate and transmit signals upon intercellular interaction, little is known about how EphB6 functions in the absence of intrinsic kinase activity. Here, we unveil a molecular mechanism of cell-cell interaction driven by EphB6. We identify ephrinB1 as a cognate ligand of EphB6 and show that in trans interaction of EphB6 with ephrinB1 on neighboring cells leads to the formation of large co-clusters at the plasma membrane...

Cell-to-cell communication through secreted Wnt ligands that bind to members of the Frizzled (Fzd) family of transmembrane receptors is critical for development and homeostasis. Wnt9a signals through Fzd9b, the co-receptor LRP5 or LRP6 (LRP5/6), and the epidermal growth factor receptor (EGFR) to promote early proliferation of zebrafish and human hematopoietic stem cells during development. Here, we developed fluorescently labeled, biologically active Wnt9a and Fzd9b fusion proteins to demonstrate that EGFR-dependent endocytosis of the ligand-receptor complex was required for signaling...

The BCR allows for Ag-driven B cell activation and subsequent Ag endocytosis, processing, and presentation to recruit T cell help. Core drivers of BCR signaling and endocytosis are motifs within the receptor's cytoplasmic tail (primarily CD79). However, BCR function can be tuned by other proximal cellular elements, such as CD20 and membrane lipid microdomains. To identify additional proteins that could modulate BCR function, we used a proximity-based biotinylation technique paired with mass spectrometry to identify molecular neighbors of the murine IgM BCR...

Antisense oligonucleotide (ASO) therapy is a novel therapeutic approach in which ASO specifically binds target mRNA, resulting in mRNA degradation; however, cellular uptake of ASOs remains critically low, warranting improvement. Transient receptor potential canonical (TRPC) channels regulate Ca2+ influx and are activated upon stimulation by phospholipase C-generated diacylglycerol. Herein, we report that a novel TRPC3/C6/C7 activator, L687, can induce cellular ASO uptake. L687-induced ASO uptake was enhanced in a dose- and incubation-time-dependent manner...

Here, we introduce a novel and effective approach utilizing a cathepsin B cleavage albumin-binding SN38 prodrug specifically designed for the treatment of metastatic breast cancer. Termed Mal-va-mac-SN38, our prodrug exhibits a unique ability to rapidly and covalently bind with endogenous albumin, resulting in the formation of HSA-va-mac-SN38. This prodrug demonstrates exceptional stability in human plasma. Importantly, HSA-va-mac-SN38 showcases an impressive enhancement in cellular uptake by 4T1 breast cancer cells, primarily facilitated through caveolin-mediated endocytosis...

Since its discovery over three decades ago, signal transducer and activator of transcription 1 (STAT1) has been extensively studied as a central mediator for interferons (IFNs) signaling and antiviral defense. Here, using genetic and biochemical assays, we unveil Thr748 as a conserved IFN-independent phosphorylation switch in Stat1, which restricts IFN signaling and promotes innate inflammatory responses following the recognition of the bacterial-derived toxin lipopolysaccharide (LPS). Genetically engineered mice expressing phospho-deficient threonine748-to-alanine (T748A) mutant Stat1 are resistant to LPS-induced lethality...

Endocytosis is a common process observed in most eukaryotic cells, although its complexity varies among different organisms. In Trypanosoma brucei , the endocytic machinery is under special selective pressure because rapid membrane recycling is essential for immune evasion. This unicellular parasite effectively removes host antibodies from its cell surface through hydrodynamic drag and fast endocytic internalization. The entire process of membrane recycling occurs exclusively through the flagellar pocket, an extracellular organelle situated at the posterior pole of the spindle-shaped cell...

The design of intracellular delivery systems for protein drugs remains a challenge due to limited delivery efficacy and serum stability. Herein, we propose a reversible assembly strategy to assemble cargo proteins and phenolic polymers into stable nanoparticles for this purpose using a heterobifunctional adaptor (2-formylbenzeneboronic acid). The adaptor is easily decorated on cargo proteins via iminoboronate chemistry and further conjugates with catechol-bearing polymers to form nanoparticles via boronate diester linkages...

PURPOSE: Hypoxia is often associated with glioma chemoresistance, and alleviating hypoxia is also crucial for improving treatment efficacy. However, although there are already some methods that can improve efficacy by alleviating hypoxia, real-time monitoring that can truly achieve hypoxia relief and efficacy feedback still needs to be explored. METHODS: AQ4N/Gd@PDA-FA nanoparticles (AGPF NPs) were synthesized using a one-pot method and were characterized. The effects of AGPF NPs on cell viability, cellular uptake, and apoptosis were investigated using the U87 cell line...

Receptor-mediated cellular uptake of specific ligands constitutes an important step in the dynamic regulation of individual protein levels in extracellular fluids. With a focus on the inflammatory lung, we here performed a proteomics-based search for novel ligands regulated by the mannose receptor (MR), a macrophage-expressed endocytic receptor. Wildtype and MR-deficient mice were exposed to lipopolysachharide (LPS), after which the protein content in their lung epithelial lining fluid (ELF) was compared by tandem mass tag-based mass spectrometry...

Triple-negative breast cancer (TNBC) is an aggressive subtype characterized by the absence of commonly targeted receptors. Unspecific chemotherapy is currently the main therapeutic option, with poor results. Another major challenge is the frequent appearance of brain metastasis (BM) associated with a significant decrease in patient overall survival. The treatment of BM is even more challenging due to the presence of the blood-brain barrier (BBB). Here, we present a dual-acting peptide (PepH3-vCPP2319) designed to tackle TNBC/BM, in which a TNBC-specific anticancer peptide (ACP) motif (vCPP2319) is joined to a BBB peptide shuttle (BBBpS) motif (PepH3)...

Alzheimer's disease (AD) pathology is characterized by amyloid beta (Aβ) plaques and dysfunctional autophagy. Aβ is generated by sequential proteolytic cleavage of amyloid precursor protein (APP), and the site of intracellular APP processing is highly debated, which may include autophagosomes. Here, we investigated the involvement of autophagy, including the role of ATG9 in APP intracellular trafficking and processing by applying the RUSH system, which allows studying the transport of fluorescently labeled mCherry-APP-EGFP in a systematic way, starting from the endoplasmic reticulum...

EH domain-containing protein 2 (EHD2) is a member of the EHD protein family and is mainly located in the plasma membrane, but can also be found in the cytoplasm and endosomes. EHD2 is also a nuclear-cytoplasmic shuttle protein. After entering the cell nuclear, EHD2 acts as a corepressor of transcription to inhibit gene transcription. EHD2 regulates a series of biological processes. As a key regulator of endocytic transport, EHD2 is involved in the formation and maintenance of endosomal tubules and vesicles, which are critical for the intracellular transport of proteins and other substances...

Nipah virus (NiV) is a highly lethal zoonotic virus with a potential large-scale outbreak, which poses a great threat to world health and security. In order to explore more potential factors associated with NiV, a proximity labeling method was applied to investigate the F, G, and host protein interactions systematically. We screened 1996 and 1524 high-confidence host proteins that interacted with the NiV fusion (F) glycoprotein and attachment (G) glycoprotein in HEK293T cells by proximity labeling technology, and 863 of them interacted with both F and G...

Adrenergic pathways represent the main channel of communication between the nervous system and the immune system. During inflammation, blood monocytes migrate within tissue and differentiate into macrophages, which polarize to M1 or M2 macrophages with tissue-damaging or -reparative properties, respectively. This study investigates whether the β-adrenergic receptor (β-AR)-blocking drug propranolol modulates the monocyte-to-macrophage differentiation process and further influences macrophages in their polarization toward M1- and M2-like phenotypes...

This first-in-its-class proof-of-concept study explored the use of bionanovesicles for the delivery of photosensitizer into cultured cholangiocarcinoma cells and subsequent treatment by photodynamic therapy (PDT). Two types of bionanovesicles were prepared: cellular vesicles (CVs) were fabricated by sonication-mediated nanosizing of cholangiocarcinoma (TFK-1) cells, whereas cell membrane vesicles (CMVs) were produced by TFK-1 cell and organelle membrane isolation and subsequent nanovesicularization by sonication...

Baicalein has been implicated in the chemotherapy overcoming triple-negative breast cancer (TNBC). However, many unanswered questions remain regarding its role in treating TNBC. Here, we sought to demonstrate the molecular pathway mediated by baicalein in TNBC. Lysine-specific demethylase 4E (KDM4E), reduced in TNBC cells, was identified as a target protein of baicalein, and baicalein enhanced the protein expression and stability of KDM4E in TNBC cells. Knockdown of KDM4E attenuated the inhibitory effect of baicalein on TNBC cell activity, as demonstrated by intensified mobility, viability, and apoptosis resistance in TNBC cells...

Autosomal dominant polycystic kidney disease (ADPKD) occurs when the proteins Polycystin-1 (PC1, PKD1 ) and Polycystin-2 (PC2, PKD2 ) contain mutations. PC1 is a large membrane receptor that can interact and form a complex with the calcium-permeable cation channel PC2. This complex localizes to the plasma membrane, primary cilia and ER. Dysregulated calcium signalling and consequential alterations in downstream signalling pathways in ADPKD are linked to cyst formation and expansion; however, it is not completely understood how PC1 and PC2 regulate calcium signalling...

BACKGROUND: Acute pancreatitis (AP) is a clinically common acute abdominal disease, whose pathogenesis remains unclear. The severe patients usually have multiple complications and lack specific drugs, leading to a high mortality and poor outcome. Acinar cells are recognized as the initial site of AP. However, there are no precise single-cell transcriptomic profiles to decipher the landscape of acinar cells during AP, which are the missing pieces of jigsaw we aimed to complete in this study...

The endothelial glycocalyx, located at the luminal surface of the endothelium, plays an important role in the regulation of leukocyte adhesion, vascular permeability, and vascular homeostasis. Endomucin (EMCN), a component of the endothelial glycocalyx, is a mucin-like transmembrane glycoprotein selectively expressed by venous and capillary endothelium. We have previously shown that knockdown of EMCN impairs retinal vascular development in vivo and vascular endothelial growth factor 165 isoform (VEGF165)-induced cell migration, proliferation, and tube formation by human retinal endothelial cells in vitro and that EMCN is essential for VEGF165-stimulated clathrin-mediated endocytosis and signaling of VEGF receptor 2 (VEGFR2)...