A chemical inhibitor of IST1-CHMP1B interaction impairs endosomal recycling and induces noncanonical LC3 lipidation.

The endosomal sorting complex required for transport (ESCRT) machinery constitutes multisubunit protein complexes that play an essential role in membrane remodeling and trafficking. ESCRTs regulate a wide array of cellular processes, including cytokinetic abscission, cargo sorting into multivesicular bodies (MVBs), membrane repair, and autophagy. Given the versatile functionality of ESCRTs, and the intricate organizational structure of the ESCRT machinery, the targeted modulation of distinct ESCRT complexes is considerably challenging.

Rho GTPase activity crosstalk mediated by Arhgef11 and Arhgef12 coordinates cell protrusion-retraction cycles.

Rho GTPases play a key role in the spatio-temporal coordination of cytoskeletal dynamics during cell migration. Here, we directly investigate crosstalk between the major Rho GTPases Rho, Rac and Cdc42 by combining rapid activity perturbation with activity measurements in mammalian cells. These studies reveal that Rac stimulates Rho activity.

ATG12-ATG5-TECPR1: an alternative E3-like complex utilized during the cellular response to lysosomal membrane damage.

ATG16L1 is an essential component of the Atg8-family protein conjugation machinery, providing membrane targeting for the ATG12-ATG5 conjugate. Recently, we identified an alternative E3-like complex that functions independently of ATG16L1. This complex utilizes the autophagosome-lysosome tethering factor TECPR1 for membrane targeting.

Inducin Triggers LC3-Lipidation and ESCRT-Mediated Lysosomal Membrane Repair.

Lipidation of the LC3 protein has frequently been employed as a marker of autophagy. However, LC3-lipidation is also triggered by stimuli not related to canonical autophagy. Therefore, characterization of the driving parameters for LC3 lipidation is crucial to understanding the biological roles of LC3.

An ATG12-ATG5-TECPR1 E3-like complex regulates unconventional LC3 lipidation at damaged lysosomes.

Lysosomal membrane damage represents a threat to cell viability. As such, cells have evolved sophisticated mechanisms to maintain lysosomal integrity. Small membrane lesions are detected and repaired by the endosomal sorting complex required for transport (ESCRT) machinery while more extensively damaged lysosomes are cleared by a galectin-dependent selective macroautophagic pathway (lysophagy).

Eating while intoxicated: characterizing the molecular mechanism behind toxin MakA-regulated autophagy.

Extracellular pathogens utilize secreted virulence factors to regulate host cell function. Recently we characterized the molecular mechanism behind host macroautophagy/autophagy regulation by the toxin MakA. Cholesterol binding at the plasma membrane induces MakA endocytosis and pH-dependent pore assembly.

V. cholerae MakA is a cholesterol-binding pore-forming toxin that induces non-canonical autophagy.

Pore-forming toxins (PFTs) are important virulence factors produced by many pathogenic bacteria. Here, we show that the Vibrio cholerae toxin MakA is a novel cholesterol-binding PFT that induces non-canonical autophagy in a pH-dependent manner. MakA specifically binds to cholesterol on the membrane at pH < 7.

Ultrafast and selective labeling of endogenous proteins using affinity-based benzotriazole chemistry.

Chemical modification of proteins is enormously useful for characterizing protein function in complex biological systems and for drug development. Selective labeling of native or endogenous proteins is challenging owing to the existence of distinct functional groups in proteins and in living systems. Chemistry for rapid and selective labeling of proteins remains in high demand.

Synthesis of 20-Membered Macrocyclic Pseudo-Natural Products Yields Inducers of LC3 Lipidation.

Design and synthesis of pseudo-natural products (PNPs) through recombination of natural product (NP) fragments in unprecedented arrangements enables the discovery of novel biologically relevant chemical matter. With a view to wider coverage of NP-inspired chemical and biological space, we describe the combination of this principle with macrocycle formation. PNP-macrocycles were synthesized efficiently in a stereoselective one-pot procedure including the 1,3-dipolar cycloadditions of different dipolarophiles with dimeric cinchona alkaloid-derived azomethine ylides formed in situ.

Imaging of Spatial Cycling of Rab GTPase in the Cell.

Rab GTPases (>60 members in human) function as master regulators of intracellular membrane trafficking. To fulfill their functions, Rab proteins need to localize on specific membranes in cells. It remains elusive how the distinct spatial distribution of Rab GTPases in the cell is regulated.

Spatiotemporal Imaging of Small GTPase Activity Using Conformational Sensors for GTPase Activity (COSGA).

Small GTPases cycle between active GTP bound and inactive GDP bound forms in live cells. They act as molecular switches and regulate diverse cellular processes at different times and locations in the cell. Spatiotemporal visualization of their activity provides important insights into dynamics of cellular signaling.

Thermal proteome profiling identifies the membrane-bound purinergic receptor P2X4 as a target of the autophagy inhibitor indophagolin.

Signaling pathways are frequently activated through signal-receiving membrane proteins, and the discovery of small molecules targeting these receptors may yield insights into their biology. However, due to their intrinsic properties, membrane protein targets often cannot be identified by means of established approaches, in particular affinity-based proteomics, calling for the exploration of new methods. Here, we report the identification of indophagolin as representative member of an indoline-based class of autophagy inhibitors through a target-agnostic phenotypic assay.

Optogenetic Tuning Reveals Rho Amplification-Dependent Dynamics of a Cell Contraction Signal Network.

Local cell contraction pulses play important roles in tissue and cell morphogenesis. Here, we improve a chemo-optogenetic approach and apply it to investigate the signal network that generates these pulses. We use these measurements to derive and parameterize a system of ordinary differential equations describing temporal signal network dynamics.

cytotoxin MakA induces noncanonical autophagy resulting in the spatial inhibition of canonical autophagy.

Autophagy plays an essential role in the defense against many microbial pathogens as a regulator of both innate and adaptive immunity. Some pathogens have evolved sophisticated mechanisms that promote their ability to evade or subvert host autophagy. Here, we describe a novel mechanism of autophagy modulation mediated by the recently discovered cytotoxin, motility-associated killing factor A (MakA).

RAB33B recruits the ATG16L1 complex to the phagophore via a noncanonical RAB binding protein.

Autophagosome formation is a fundamental process in macroautophagy/autophagy, a conserved self-eating mechanism in all eukaryotes, which requires the conjugating ATG (autophagy related) protein complex, ATG12-ATG5-ATG16L1 and lipidated MAP1LC3/LC3 (microtubule associated protein 1 light chain 3). How the ATG12-ATG5-ATG16L1 complex is recruited to membranes is not fully understood. Here, we demonstrated that RAB33B plays a key role in recruiting the ATG16L1 complex to phagophores during starvation-induced autophagy.

Factors associated with spatial distribution of severe fever with thrombocytopenia syndrome.

Severe fever with thrombocytopenia syndrome (SFTS) was firstly identified in mainland China in 2009 and the geographic distribution has expanded in recent years. In this study, we constructed ecological niche models (ENM) of SFTS with meteorological factors, environmental factor, and density of domestic animals using MaxEnt. We found four significant associated factors including altitude, yearly average temperature, yearly accumulated precipitation, and yearly average relative humidity which accounted for 94.

Distinct Mechanisms for Processing Autophagy Protein LC3-PE by RavZ and ATG4B.

Autophagy is a conserved catabolic process involved in the elimination of proteins, organelles and pathogens in eukaryotic cells. Lipidated LC3 proteins that are conjugated to phosphatidylethanolamine (PE) play a key role in autophagosome biogenesis. Endogenous ATG4-mediated deconjugation of LC3-PE is required for LC3 recycling.

Phenotyping Reveals Targets of a Pseudo-Natural-Product Autophagy Inhibitor.

Pseudo-natural-product (NP) design combines natural product fragments to provide unprecedented NP-inspired compounds not accessible by biosynthesis, but endowed with biological relevance. Since the bioactivity of pseudo-NPs may be unprecedented or unexpected, they are best evaluated in target agnostic cell-based assays monitoring entire cellular programs or complex phenotypes. Here, the Cinchona alkaloid scaffold was merged with the indole ring system to synthesize indocinchona alkaloids by Pd-catalyzed annulation.

Inhibition of K-Ras4B-plasma membrane association with a membrane microdomain-targeting peptide.

The association of K-Ras4B protein with plasma membrane (PM) is required for its signaling activity. Thus, direct inhibition of K-Ras4B-PM interaction could be a potential anti-Ras therapeutic strategy. However, it remains challenging to modulate such protein-PM interaction.

Patient with severe fever with thrombocytopenia syndrome virus infection and central nervous system disturbance in Dongyang, Zhejiang Province, China, 2017.

Background: Severe fever with thrombocytopenia syndrome (SFTS) is an emerging hemorrhagic fever that was first described in China in 2011. We report a patient who died of Severe fever with thrombocytopenia syndrome virus (SFTSV) infection, with a rapidly progressive central nervous system (CNS) disturbance, in Dongyang, Zhejiang Province, China, in 2017.

Case Presentation: A 64-year-old man was admitted to hospital after 4 days of fever.

Toward the role of cholesterol and cholesterol transfer protein in autophagosome biogenesis.

A forward chemical genetic approach led to identification of autogramins as novel autophagy inhibitors. Autogramins selectively target the cholesterol transfer protein GRAMD1A (GRAM domain containing 1A). Autogramins compete with cholesterol binding to the StART domain of GRAMD1A, thereby inhibiting its cholesterol transfer activity.

Light-Induced Dimerization Approaches to Control Cellular Processes.

Light-inducible approaches provide a means to control biological systems with spatial and temporal resolution that is unmatched by traditional genetic perturbations. Recent developments of optogenetic and chemo-optogenetic systems for induced proximity in cells facilitate rapid and reversible manipulation of highly dynamic cellular processes and have become valuable tools in diverse biological applications. New expansions of the toolbox facilitate control of signal transduction, genome editing, "painting" patterns of active molecules onto cellular membranes, and light-induced cell cycle control.

The cholesterol transfer protein GRAMD1A regulates autophagosome biogenesis.

Autophagy mediates the degradation of damaged proteins, organelles and pathogens, and plays a key role in health and disease. Thus, the identification of new mechanisms involved in the regulation of autophagy is of major interest. In particular, little is known about the role of lipids and lipid-binding proteins in the early steps of autophagosome biogenesis.