Toward decoding spatiotemporal signaling activities of reactive immunometabolites with precision immuno-chemical biology tools.

Immune-cell reprogramming driven by mitochondria-derived reactive electrophilic immunometabolites (mt-REMs-e.g., fumarate, itaconate) is an emerging phenomenon of major biomedical importance.

Monitoring On-Target Signaling Responses in Larval Zebrafish - Z-REX Unmasks Precise Mechanisms of Electrophilic Drugs and Metabolites.

Reactive metabolites and related electrophilic drugs are among the most challenging small molecules to study. Conventional approaches to deconstruct the mode of action (MOA) of such molecules leverage bulk treatment of experimental specimens with an excess of a specific reactive species. In this approach, the high reactivity of electrophiles renders non-discriminate labeling of the proteome in a time- and context-dependent manner; redox-sensitive proteins and processes can also be indirectly and often irreversibly affected.

Z-REX: shepherding reactive electrophiles to specific proteins expressed tissue specifically or ubiquitously, and recording the resultant functional electrophile-induced redox responses in larval fish.

This Protocol Extension describes the adaptation of an existing Protocol detailing the use of targetable reactive electrophiles and oxidants, an on-demand redox targeting toolset in cultured cells. The adaptation described here is for use of reactive electrophiles and oxidants technologies in live zebrafish embryos (Z-REX). Zebrafish embryos expressing a Halo-tagged protein of interest (POI)-either ubiquitously or tissue specifically-are treated with a HaloTag-specific small-molecule probe housing a photocaged reactive electrophile (either natural electrophiles or synthetic electrophilic drug-like fragments).

Z-REX uncovers a bifurcation in function of Keap1 paralogs.

Studying electrophile signaling is marred by difficulties in parsing changes in pathway flux attributable to on-target, vis-à-vis off-target, modifications. By combining bolus dosing, knockdown, and Z-REX-a tool investigating on-target/on-pathway electrophile signaling, we document that electrophile labeling of one zebrafish-Keap1-paralog (zKeap1b) stimulates Nrf2- driven antioxidant response (AR) signaling (like the human-ortholog). Conversely, zKeap1a is a dominant-negative regulator of electrophile-promoted Nrf2-signaling, and itself is nonpermissive for electrophile-induced Nrf2-upregulation.

Investigating a Boronate-Affinity-Guided Acylation Reaction for Labelling Native Antibodies.

The excellent molecular recognition capabilities of monoclonal antibodies (mAbs) have opened up exciting opportunities for biotherapeutic discovery. Taking advantage of the full potential of this tool necessitates affinity ligands capable of conjugating directly with small molecules to a defined degree of biorthogonality, especially when modifying natural Abs. Herein, a bioorthogonal boronate-affinity-based Ab ligand featuring a 4-(dimethylamino)pyridine and an S-aryl thioester to label full-length Abs is reported.

The not so identical twins: (dis)similarities between reactive electrophile and oxidant sensing and signaling.

In this tutorial review, we compare and contrast the chemical mechanisms of electrophile/oxidant sensing, and the molecular mechanisms of signal propagation. We critically analyze biological systems in which these different pathways are believed to be manifest and what the data really mean. Finally, we discuss applications of this knowledge to disease treatment and drug development.

Wdr1 and cofilin are necessary mediators of immune-cell-specific apoptosis triggered by Tecfidera.

Despite the emerging importance of reactive electrophilic drugs, deconvolution of their principal targets remains difficult. The lack of genetic tractability/interventions and reliance on secondary validation using other non-specific compounds frequently complicate the earmarking of individual binders as functionally- or phenotypically-sufficient pathway regulators. Using a redox-targeting approach to interrogate how on-target binding of pleiotropic electrophiles translates to a phenotypic output in vivo, we here systematically track the molecular components attributable to innate immune cell toxicity of the electrophilic-drug dimethyl fumarate (Tecfidera®).

Regioselective S2-Type Reaction for the Oriented and Irreversible Immobilization of Antibodies to a Glass Surface Assisted by Boronate Formation.

Antibodies have exquisite specificities for molecular recognition, which have led to their incorporation into array sensors that are crucial for research, diagnostic, and therapeutic applications. Many of these platforms rely heavily on surface-bound reactive groups to covalently tether antibodies to solid substrates; however, this strategy is hindered by a lack of orientation control over antibody immobilization. Here, we report a mild electrophilic phenylsulfonate (tosylate) ester-containing boronic acid affinity ligand for attaching antibodies to glass slides.

Real-Time Identification of Irrigation Water Pollution Sources and Pathways with a Wireless Sensor Network and Blockchain Framework.

Real-time identification of irrigation water pollution sources and pathways (PSP) is crucial to ensure both environmental and food safety. This study uses an integrated framework based on the Internet of Things (IoT) and the blockchain technology that incorporates a directed acyclic graph (DAG)-configured wireless sensor network (WSN), and GIS tools for real-time water pollution source tracing. Water quality sensors were installed at monitoring stations in irrigation channel systems within the study area.