The crustacean DNA virus tegument protein VP26 binds to SNAP29 to inhibit SNARE complex assembly and autophagic degradation.

Autophagy generally functions as a cellular surveillance mechanism to combat invading viruses, but viruses have evolved various strategies to block autophagic degradation and even subvert it to promote viral propagation. White spot syndrome virus (WSSV) is the most highly pathogenic crustacean virus, but little is currently known about whether crustacean viruses such as WSSV can subvert autophagic degradation for escape. Here, we show that even though WSSV proliferation triggers the accumulation of autophagosomes, autophagic degradation is blocked in the crustacean species red claw crayfish. Interestingly, the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex including CqSNAP29, CqVAMP7, and the novel autophagosome SNARE protein CqSyx12 is required for autophagic flux to restrict WSSV replication, as revealed by gene silencing experiments. Simultaneously, the expressed WSSV tegument protein VP26, which likely localizes on autophagic membrane mediated by its transmembrane region, binds the Qb-SNARE domain of CqSNAP29 to competitively inhibit the binding of CqSyx12-Qa-SNARE with CqSNAP29-Qb-SNARE; this in turn disrupts the assembly of the CqSyx12-SNAP29-VAMP7 SNARE complex, which is indispensable for the proposed fusion of autophagosomes and lysosomes. Consequently, the autophagic degradation of WSSV is likely suppressed by the expressed VP26 protein in vivo in crayfish, thus probably protecting WSSV components from degradation via the autophagosome-lysosome pathway, resulting in evasion by WSSV. Collectively, these findings highlight how a DNA virus can subvert autophagic degradation by impairing the assembly of the SNARE complex to achieve evasion, paving the way for understanding host-DNA virus interactions from an evolutionary point of view, from crustaceans to mammals.IMPORTANCEWhite spot syndrome virus (WSSV) is one of the largest animal DNA viruses in terms of its genome size and has caused huge economic losses in the farming of crustaceans such as shrimp and crayfish. Detailed knowledge of WSSV-host interactions is still lacking, particularly regarding viral escape from host immune clearance. Intriguingly, we found that the presence of WSSV-VP26 might inhibit the autophagic degradation of WSSV in vivo in the crustacean species red claw crayfish. Importantly, this study is the first to show that viral protein VP26 functions as a core factor to benefit WSSV escape by disrupting the assembly of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex, which is necessary for the proposed fusion of autophagosomes with lysosomes for subsequent degradation. These findings highlight a novel mechanism of DNA virus evasion by blocking SNARE complex assembly and identify viral VP26 as a key candidate for anti-WSSV targeting.

Fluorinated Nonporous Adaptive Cages for the Efficient Removal of Perfluorooctanoic Acid from Aqueous Source Phases.

Per- and polyfluoroalkyl substances (PFAS) accumulate in water resources and pose serious environmental and health threats due to their nonbiodegradable nature and long environmental persistence times. Strategies for the efficient removal of PFAS from contaminated water are needed to address this concern. Here, we report a fluorinated nonporous adaptive crystalline cage (F-Cage 2) that exploits electrostatic interaction, hydrogen bonding, and F-F interactions to achieve the efficient removal of perfluorooctanoic acid (PFOA) from aqueous source phases. F-Cage 2 exhibits a high second-order kobs value of approximately 441,000 g mg-1 h-1 for PFOA and a maximum PFOA adsorption capacity of 45 mg g-1. F-Cage 2 can decrease PFOA concentrations from 1500 to 6 ng L-1 through three rounds of flow-through purification, conducted at a flow rate of 40 mL h-1. Elimination of PFOA from PFOA-loaded F-Cage 2 is readily achieved by rinsing with a mixture of MeOH and saturated NaCl. Heating at 80 °C under vacuum then makes F-Cage 2 ready for reuse, as demonstrated across five successive uptake and release cycles. This work thus highlights the potential utility of suitably designed nonporous adaptive crystals as platforms for PFAS remediation.

The Combination of Quantitative Proteomics and Systems Genetics Analysis Reveals that PTN Is Associated with Sleep-Loss-Induced Cognitive Impairment.

Sleep loss is associated with cognitive dysfunction. However, the detailed mechanisms remain unclear. In this study, we established a para-chlorophenylalanine (PCPA)-induced insomniac mouse model with impaired cognitive function. Mass-spectrometry-based proteomics showed that the expression of 164 proteins was significantly altered in the hippocampus of the PCPA mice. To identify critical regulators among the potential markers, a transcriptome-wide association screening was performed in the BXD mice panel. Among the candidates, the expression of pleiotrophin (Ptn) was significantly associated with cognitive functions, indicating that Ptn-mediates sleep-loss-induced cognitive impairment. Gene co-expression analysis further revealed the potential mechanism by which Ptn mediates insomnia-induced cognitive impairment via the MAPK signaling pathway; that is, the decreased secretion of Ptn induced by insomnia leads to reduced binding to Ptprz1 on the postsynaptic membrane with the activation of the MAPK pathway via Fos and Nr4a1, further leading to the apoptosis of neurons. In addition, Ptn is genetically trans-regulated in the mouse hippocampus and implicated in neurodegenerative diseases in human genome-wide association studies. Our study provides a novel biomarker for insomnia-induced cognitive impairment and a new strategy for seeking neurological biomarkers by the integration of proteomics and systems genetics.

Silencing DTX3L Inhibits the Progression of Cervical Carcinoma by Regulating PI3K/AKT/mTOR Signaling Pathway.

Cervical carcinoma (CC) is the second most prevalent gynecologic cancer in females across the world. To obtain a better understanding of the mechanisms underlying the development of CC, high-resolution label-free mass spectrometry was performed on CC and adjacent normal tissues from eight patients. A total of 2631 proteins were identified, and 46 significant differently expressed proteins (DEPs) were found between CC and normal tissues (p < 0.01, fold change >10 or <0.1). Ingenuity pathway analysis revealed that the majority of the proteins were involved in the regulation of eIF4 and p70S6K signaling and mTOR signaling. Among 46 DEPs, Integrinß6 (ITGB6), PPP1CB, TMPO, PTGES3 (P23) and DTX3L were significantly upregulated, while Desmin (DES) was significantly downregulated in CC tissues compared with the adjacent normal tissues. In in vivo and in vitro experiments, DTX3L knockdown suppressed CC cell proliferation, migration, invasion and xenograft tumorigenesis, and enhanced cell apoptosis. Combination of silencing DTX3L and cisplatin treatment induced higher apoptosis percentage compared to cisplatin treatment alone. Moreover, DTX3L silencing inhibited the PI3K/AKT/mTOR signal pathway. Thus, our results suggested DTX3L could regulate CC progression through the PI3K/AKT/mTOR signal pathway and is potentially a novel biomarker and therapeutic target for CC.

Reversible Iodine Capture by Nonporous Adaptive Crystals of a Bipyridine Cage.

The ability to capture radioactive iodine species is crucial for nuclear accident preparedness and nuclear waste treatment; however, it remains a challenge. Here we report a new readily obtainable nitrogen-rich nonporous cage (BPy-Cage) based on bipyridine building blocks that supports iodine capture. This cage is able to capture not only volatile iodine in vapor form but also iodine dissolved in various organic solvents or aqueous media with an iodine uptake capacity of up to 3.23 g g-1. The iodine within the cage (I2@BPy-Cage) can be released quickly upon immersing the bound solid form in DMF, allowing for control over acylation reactions. The cage solids reported here could be reused several times without substantial loss in their iodine capture performance. The effectiveness of the present system is ascribed to its ability to support strong iodine-bipyridine nitrogen lone pair interactions.

Calix[4]pyrrole-Based Azo-Bridged Porous Organic Polymer for Bromine Capture.

The toxicity, corrosiveness, and volatility of elemental bromine presents challenges for its safe storage and transportation. Purification from other halogens is also difficult. Here, we report an easy-to-prepare calix[4]pyrrole-based azo-bridged porous organic polymer (C4P-POP) that supports efficient bromine capture. C4P-POP was found to capture bromine as a vapor and from a cyclohexane source phase with maximum uptake capacities of 3.6 and 3.4 g·g-1, respectively. Flow-through adsorption experiments revealed that C4P-POP removes 80% of the bromine from a 4.0 mM cyclohexane solution at a flow rate of 45 mL·h-1. C4P-POP also allowed the selective capture of bromine from a 1:1 mixture of bromine and iodine in cyclohexane.

GAD1 alleviates injury-induced optic neurodegeneration by inhibiting retinal ganglion cell apoptosis.

The degeneration of the optic nerve narrows the visual field, eventually causing overall vision loss. This study aimed to identify global protein changes in the retina of optic nerve crushing (ONC) mice and to identify key regulators and pathways involved in injury-induced cell death during the progression of optic neurodegeneration. Label-free quantitative proteomics combined with bioinformatic analysis was performed on retinal protein extracts from ONC and sham-operated mice. Among the 1433 proteins detected, 121 proteins were differentially expressed in the retina of ONC mice. Further bioinformatic analysis showed that various metabolic pathways, including glutamate metabolism and γ-aminobutyric acid (GABA) synthesis, were significantly dysregulated in the injured mouse retinas. Glutamate decarboxylase 1 (GAD1) is the enzyme that converts glutamate into GABA, which was significantly up-regulated during ONC injury. Exogenous GAD1 treatment increased retinal ganglion cell (RGC) survival in the ONC-injured retina. In addition, changes in GAD1 expression were also observed in several other ophthalmic diseases. Vascular endothelial growth factor B (VEGF-B) has previously been reported to protect RGCs from apoptosis and positively regulated the expression of GAD1 in the retina. Notably, combination treatment with GAD1 and VEGF-B also provided strong protection against injury-induced RGC apoptosis. These results suggest that GAD1 expression may serve as an intrinsic protective mechanism that is commonly activated during retinal injury. Targeting GAD1 may serve as a potential strategy to treat optic neurodegenerative diseases.

Nonporous Adaptive Calix[4]pyrrole Crystals for Polar Compound Separations.

The use of molecular crystalline materials for the separation and purification of chemical raw materials, particularly polar compounds with similar physical and chemical properties, represents an ongoing challenge. This is particularly true for volatile feedstocks that form binary azeotropes. Here we report a new cavity-extended version of calix[4]pyrrole (C4P) that readily forms nonporous adaptive crystals (NACs). These C4P-based NACs allow pyridine to be separated from toluene/pyridine mixtures with nearly 100% purity, as well as the removal of 1,4-dioxane from 1,4-dioxane/water mixtures with high adsorption capacity. Removal of the polar guest (pyridine or 1,4-dioxane) from the guest-loaded NACs by heating under vacuum produces the guest-free crystalline form. In the case of both guests, the C4P material could be reused as demonstrated through 10 uptake and release cycles without apparent performance loss.

"Texas-Sized" Molecular Boxes: From Chemistry to Applications.

The design and synthesis of novel macrocyclic host molecules continues to attract attention because such species play important roles in supramolecular chemistry. However, the discovery of new classes of macrocycles presents a considerable challenge due to the need to embody by design effective molecular recognition features, as well as ideally the development of synthetic routes that permit further functionalization. In 2010, we reported a new class of macrocyclic hosts: a set of tetracationic imidazolium macrocycles, which we termed "Texas-sized" molecular boxes (TxSBs) in homage to Stoddart's classic "blue box" (CBPQT4+). Compared with the rigid blue box, the first generation TxSB displayed considerably greater conformational flexibility and a relatively large central cavity, making it a good host for a variety of electron-rich guests. In this review, we provide a comprehensive summary of TxSB chemistry, detailing our recent progress in the area of anion-responsive supramolecular self-assembly and applications of the underlying chemistry to water purification, information storage, and controlled drug release. Our objective is to provide not only a review of the fundamental findings, but also to outline future research directions where TxSBs and their constructs may have a role to play.

Soft Materials Constructed Using Calix[4]pyrrole- and "Texas-Sized" Box-Based Anion Receptors.

Soft materials have received considerable attention from supramolecular chemists and material scientists alike. This interest reflects the advantages provided by their soft, flexible nature and the convenience of the molecular self-assembly that underlies their preparation. Common soft supramolecular materials include polymeric gels, supramolecular polymers, nanoaggregates, and membranes. Polymeric gels are solidlike networks of cross-linked polymer chains. Supramolecular polymers contain repeat units connected through reversible non-covalent bonds. Nanoaggregates are formed as a result of hydrophobic interactions involving amphiphilic building blocks. Because of the presence of non-covalent interactions, supramolecular soft materials typically display stimuli-responsive or adaptive features. Various macrocyclic hosts, such as cyclodextrins, crown ethers, calixarenes, cucurbiturils, and pillararenes, and many classic non-covalent interactions have been harnessed to construct supramolecular soft materials. Only recently has anion binding been used as the underlying recognition motif. Anions are ubiquitous in the natural world. Their importance has inspired efforts to achieve good anion binding and to exploit anion recognition in a number of fields, including extraction, transport, sensing, and catalysis. Most of this effort has involved the use of stand-alone anion receptors. On the other hand, soft materials with anion recognition features could lead to new macromolecular systems of interest in the context of many application areas. In this Account, we summarize the latest efforts from our laboratory to prepare supramolecular soft materials, including polymeric gels, supramolecular polymers, and nanoaggregates, with bona fide anion recognition features. Two anion receptor systems, namely, calix[4]pyrroles (C4Ps) and a tetraimidazolium macrocycle known as the "Texas-sized" molecular box (TxSB), have been used for this purpose. To date, TxSB-based hydrogels have been utilized to capture anions from water and for coded information applications; C4P-based organic polymeric gels have been used to extract dianions from aqueous source phases and for the on-site detection of chloride anions. Polymers containing C4P and TxSB anion recognition subunits typically display responsive features and can be modified through application of appropriately chosen external stimuli. For instance, nanoaggregates may be formed as a result of the hydrophobic interactions of C4P- and TxSB-based amphiphiles. The resulting aggregates were found to mimic the structural evolution of organelles and could be used as effective anion and ion pair extractants. This Account summarizes progress to date while underscoring potential opportunities associated with combining anion recognition and soft materials chemistry. The hope is to stimulate further advances in broad areas, including polymer science, supramolecular chemistry, biology, materials research, and information storage.

Cation-based Structural Tuning of Pyridine Dipyrrolate Cages and Morphological Control over Their Self-assembly.

Different pyridine dipyrrolate cages including cage-based dimers and polymers may be fabricated in a controlled manner from the same two starting materials, namely, an angular ligand 1 and Zn(acac)2, by changing the counter cation source. With tetrabutylammonium (TBA+) and dimethyl viologen (DMV2+), Cage-3 and Cage-5 are produced. In these cages, two ligands act as bridges and serve to connect together two cage subunits to produce higher order ensembles. In Cage-3 and Cage-5, the TBA+ and DMV2+ counter cations lie outside the cavities of the respective cages. This stands in contrast to what is seen with a previously reported system, Cage-1, wherein dimethylammonium (DMA+) counter cations reside within the cage cavity. When the counter cations are tetraethylammonium (TEA+) and bis(cyclopentadienyl) cobalt(III) (Cp2Co+), polymeric cage materials, PC-1 and PC-2, are formed, respectively. The counter cations thus serve not only to balance charge but also to tune the structural features as a whole. The organic cations used in the present study also act to modulate the further assembly of individual cages. The present cation-based tuning emerges as a new method for a fine-tuning of the multidimensional morphology of self-assembled inorganic materials.

Azobenzene-Bridged Expanded "Texas-sized" Box: A Dual-Responsive Receptor for Aryl Dianion Encapsulation.

We report an expanded "Texas-sized" molecular box (AzoTxSB) that incorporates photoresponsive azobenzene bridging subunits and anion recognition motifs. The shape of this box can be switched through light induced E ↔ Z photoisomerization of the constituent azobenzenes. This allows various anionic substrates to be bound and released by using different forms of the box. Control can also be achieved using other environmental stimuli, such as pH and anion competition.

Molecular Cursor Caliper: A Fluorescent Sensor for Dicarboxylate Dianions.

We report here the fluorescent sensing of both aromatic and linear saturated dicarboxylate anions (DC2-) (as their tetrabutylammonium salts) with different lengths and shapes in acetonitrile using a single fluorescent probe, i.e., the bis-calix[4]pyrrole-appended 9,14-diphenyl-9,14-dihydrodibenzo[a,c]phenazine (DPAC-bisC4P) incorporating a vibration-induced emission (VIE) phenazine core. Fluorescence titration studies revealed that treating DPAC-bisC4P with dicarboxylate guests capable of forming pseudomacrocyclic host-guest complexes via multiple hydrogen-bonding interactions between the dicarboxylates and calix[4]pyrrole moieties led to a blue-shift in the emission of the phenazine core. The binding-based fluorescence-tuning features of DPAC-bisC4P allow the underlying binding events and inferred structural changes to be monitored in the form of different chromaticity outputs. The analyte-induced differences in the fluorescence response to DC2- cover a wide range within the chromaticity diagram and can be visualized readily. The present system thus functions as a rudimentary dicarboxylate anion sensor. It highlights the potential benefits associated with combining a tunable VIE core with noncovalent binding interactions and thus sets the stage for the development of new fluorescent chemosensors where a single chemical entity responds to different analytes with a high level of tunability.

Platinum(IV)-Ferrocene Conjugates and Their Cyclodextrin Host-Guest Complexes.

Reported here are new platinum(IV) (Pt(IV)) complexes bearing ferrocene (Fc) moieties. These systems differ from one another only by the nature of the functional group (ester vs amide) connecting the linker to the Fc subunits. This minor structural variation (one atom difference) leads to major differences in solubility, stability, and antiproliferative activity against lung (A549) cancer cells. The host-guest chemistry of these complexes was investigated in an aqueous medium in the presence of ß-cyclodextrins (ß-CD), either free or in the form of a covalently linked Fc-Pt-ß-CD hybrid. An inclusion complex between Fc and ß-CD is formed in aqueous media, presumably as a result of hydrophobic interactions involving the Fc and the inner ß-CD cavity. Consequently, it proved possible to use a ß-CD-based strategy to purify the Pt-Fc conjugates in this study under aqueous conditions (by means of C18 silica gel columns). The use of a ß-CD adjuvant also allowed dimethyl sulfoxide (DMSO) to be avoided as an organic cosolvent in cell studies. The amide version reported here (2) proved to be more soluble, more stable, and more active than the ester analogue (11) in A549 cells. The use of a ß-CD functionalized with a fluorescent probe allowed intracellular Pt-Fc localization to be visualized by confocal fluorescence microscopy.

Supramolecular Properties of a Monocarboxylic Acid-Functionalized "Texas-Sized" Molecular Box.

A new carboxylic acid-functionalized "Texas-sized" molecular box TxSB-CO2H has been prepared by combining two separate building blocks via an iodide-catalyzed macrocyclization reaction. A single-crystal X-ray diffraction analysis revealed a paired "clip-like" dimer in the solid state. Concentration-dependent behavior is seen for samples of TxSB-CO2H as prepared, as inferred from 1H NMR spectroscopic studies carried out in DMSO- d6. However, in the presence of excess acid (1% by weight of deuterated trifluoracetic acid; TFA- d1), little evidence of aggregation is seen in DMSO- d6 except at the highest accessible concentrations. In contrast, the conjugate base form, TxSB-CO2-, produced in situ via the addition of excess triethylamine to DMSO- d6 solutions of TxSB-CO2H acts as a self-complementary monomer that undergoes self-assembly to stabilize a formal oligomer ([TxSB-CO2-] n) with a degree of polymerization of approximately 5-6 at a concentration of 70 mM. Evidence in support of the proposed oligomerization of TxSB-CO2- in solution and in the solid state came from one- and two-dimensional 1H NMR spectroscopy, X-ray crystallography, dynamic light scattering (DLS), and scanning electron microscopy (SEM). A series of solution-based analyses carried out in DMSO and DMSO- d6 provide support for the notion that the self-assembled constructs produced from TxSB-CO2- are responsive to environmental stimuli, including exposure to the acetate anion (as its tetrabutylammonium, TBA+, salt), and changes in overall concentration, temperature, and protonation state. The resulting transformations are thought to reflect the reversible nature of the underlying noncovalent interactions. They also permit the stepwise interconversion between TxSB-CO2H and [TxSB-CO2-] n via the sequential addition of triethylamine and TFA- d1. The present work thus serves to illustrate how appropriately functionalized molecular box-type macrocycles may be used to develop versatile stimuli-responsive materials. It also highlights how aggregated forms seen in the solid state are not necessarily retained under competitive solution-phase conditions.

Controlling Structure Beyond the Initial Coordination Sphere: Complexation-Induced Reversed Micelle Formation in Calix[4]pyrrole-Containing Diblock Copolymers.

A diblock copolymer containing a strapped calix[4]pyrrole-based ion pair recognition subunit has been synthesized via RAFT polymerization. As prepared, the polymer is hydrophobic and devoid of any particular morphological form. However, upon ion pair complexation, the copolymer self-assembles to generate reverse micelles in organic media. The reverse micelles formed in this way may be used to extract alkali cation and cesium halide anion salts from an aqueous source into an organic receiving phase. The polymer proved more effective as an extractant than the corresponding free ion pair receptor.

Molecular Recognition Under Interfacial Conditions: Calix[4]pyrrole-Based Cross-linkable Micelles for Ion Pair Extraction.

An anthracene-functionalized, long-tailed calix[4]pyrrole 1, containing both an anion-recognition site and cation-recognition functionality, has been synthesized and fully characterized. Upon ion pair complexation with FeF2, receptor 1 self-assembles into multimicelles in aqueous media. This aggregation process is ascribed to a change in polarity from nonpolar to amphiphilic induced upon concurrent anion and cation complexation and permits molecular recognition-based control over chemical morphology under interfacial conditions. Photoirradiation of the micelles serves to cross-link the anthracene units thus stabilizing the aggregates. The combination of ion pair recognition, micelle formation, and cross-linking can be used to extract FeF2 ion pairs from bulk aqueous solutions. The present work helps illustrate how molecular recognition and self-assembly may be used to control the chemistry of extractants at interfaces.

A Multiresponsive Amphiphilic Supramolecular Diblock Copolymer Based on Pillar[10]arene/Paraquat Complexation for Rate-Tunable Controlled Release.

Herein, for rate-tunable controlled release, the authors report a new facile method to prepare multiresponsive amphiphilic supramolecular diblock copolymers via the cooperative complexation between a water-soluble pillar[10]arene and paraquat-containing polymers in water. This supramolecular diblock copolymer can self-assemble into multiresponsive polymeric micelles at room temperature in water. The resultant micelles can be further used in the controlled release of small molecules with tunable release rates depending on the type of single stimulus and the combination of various stimuli.

A Dual-Thermoresponsive Gemini-Type Supra-amphiphilic Macromolecular [3]Pseudorotaxane Based on Pillar[10]arene/Paraquat Cooperative Complexation.

Herein, first we report the preparation of a thermoresponsive [3]pseudorotaxane from cooperative complexation between a water-soluble pillar[10]arene and a paraquat derivative in water. Then we successfully construct the first pillararene-based gemini-type supra-amphiphilic [3]pseudorotaxane from the water-soluble pillar[10]arene and a paraquat-containing poly(N-isopropylacrylamide) based on this new molecular recognition motif in water. This macromolecular [3]pseudorotaxane shows unique dual-thermoresponsiveness. Furthermore, it can self-assemble into polymeric vesicles at 37 °C in water. These vesicles can be further used in the controlled release of small molecules induced by cooling to 25 °C or heating to 60 °C.

A Dual-Responsive Bola-Type Supra-amphiphile Constructed from a Water-Soluble Calix[4]pyrrole and a Tetraphenylethene-Containing Pyridine Bis-N-oxide.

Complexation between a water-soluble calix[4]pyrrole and a ditopic pyridine N-oxide derivative in aqueous media produces a bola-type supra-amphiphile that self-assembles to produce higher order morphologies, including multilamellar vesicles and micelles depending on the pH. The present bola-type supra-amphiphile exhibits strong fluorescence due to structural changes and aggregation induced by host-guest complexation. The resulting structures may be used to recognize, encapsulate, and release non-fluorescent, water-soluble small molecules.