A high throughput assay for phosphoribosylformylglycinamidine synthase.

Metabolic reprogramming of purine biosynthesis is a hallmark of cancer metabolism and represents a critical vulnerability. The enzyme phosphoribosylformylglycinamidine synthase (PFAS) catalyzes the fourth step in de novo purine biosynthesis and has been demonstrated to be prognostic for survival of liver cancer. Despite the importance of this protein as a drug target, there are no known specific inhibitors of PFAS activity.

Efficient gene deletion of Integrin alpha 4 in primary mouse CD4 T cells using CRISPR RNA pair-mediated fragmentation.

The functional specialization of CD4 T lymphocytes into various subtypes, including T1 and T cells, is crucial for effective immune responses. T cells facilitate B cell differentiation within germinal centers, while T1 cells are vital for cell-mediated immunity against intracellular pathogens. Integrin α4, a cell surface adhesion molecule, plays significant roles in cell migration and co-stimulatory signaling.

In vivo evolution of env in SHIV-AD8-infected rhesus macaques after AAV-vectored delivery of eCD4-Ig.

eCD4-immunoglobulin (Ig) is an HIV entry inhibitor that mimics the engagement of both CD4 and CCR5 with the HIV envelope (Env) protein, a property that imbues it with remarkable potency and breadth. However, env is exceptionally genetically malleable and can evolve to escape a wide variety of entry inhibitors. Here we document the evolution of partial eCD4-Ig resistance in SHIV-AD8-infected rhesus macaques (RMs) treated with adeno-associated virus vectors encoding eCD4-Ig.

In vivo affinity maturation of the CD4 domains of an HIV-1-entry inhibitor.

Human proteins repurposed as biologics for clinical use have been engineered through in vitro techniques that improve the affinity of the biologics for their ligands. However, the techniques do not select against properties, such as protease sensitivity or self-reactivity, that impair the biologics' clinical efficacy. Here we show that the B-cell receptors of primary murine B cells can be engineered to affinity mature in vivo the human CD4 domains of the HIV-1-entry inhibitor CD4 immunoadhesin (CD4-Ig).

Delphi-driven consensus definition for mesenchymal stromal cells and clinical reporting guidelines for mesenchymal stromal cell-based therapeutics.

Background Aims: Despite promising results in pre-clinical studies, mesenchymal stromal cells (MSCs) face significant challenges in clinical translation. A scoping review by our group highlighted two key issues contributing to this gap: (i) lack of a clear and consensus definition for MSCs and (ii) under-reporting of critical parameters in MSC clinical studies. To address these issues, we conducted a modified Delphi study to establish and implement a consensus definition for MSCs and develop reporting guidelines for MSC clinical studies.

Probing a distinct druggable tubulin binding site with gatorbulins 1-7, their metabolic and physicochemical properties, and pharmacological consequences.

Microtubules, consisting of α/β-tubulin heterodimers, are prime targets for anticancer drug discovery. Gatorbulin-1 (GB1, 1a) is a recently described marine natural product that targets tubulin at a new, seventh pharmacological site at the tubulin intradimer interface. Using our previously developed robust route towards GB1 (1a), we synthesized simplified, first-generation gatorbulins, GB2-7 (1b-1g) of this highly modified cyclodepsipeptide (GB1) that does not contain any proteinogenic amino acid.

SR9883 is a novel small-molecule enhancer of α4β2* nicotinic acetylcholine receptor signaling that decreases intravenous nicotine self-administration in rats.

Background: Most smokers attempting to quit will quickly relapse to tobacco use even when treated with the most efficacious smoking cessation agents currently available. This highlights the need to develop effective new smoking cessation medications. Evidence suggests that positive allosteric modulators (PAM) and other enhancers of nicotinic acetylcholine receptor (nAChR) signaling could have therapeutic utility as smoking cessation agents.

Solid-Phase Synthesis of Diverse Macrocycles by Regiospecific 2-Pyridone Formation: Scope and Applications.

This study introduces a novel solid-phase macrocyclization method generating 2-pyridone rings. This method relies on the intramolecular condensation between secondary and tertiary dimethoxy-propionic amide units. This selective reaction leads to the formation of a single well-defined regioisomer.

Quantifying Forms and Functions of Enterohepatic Bile Acid Pools in Mice.

Backgrounds & Aims: Bile acids (BAs) are core gastrointestinal metabolites with dual functions in lipid absorption and cell signaling. BAs circulate between the liver and distal small intestine (i.e.

Bis-sulfonamido-2-phenylbenzoxazoles Validate the GroES/EL Chaperone System as a Viable Antibiotic Target.

We recently reported on small-molecule inhibitors of the GroES/GroEL chaperone system as potential antibiotics against and the ESKAPE pathogens but were unable to establish GroES/GroEL as the cellular target, leading to cell death. In this study, using two of our most potent -sulfonamido-2-phenylbenzoxazoles (PBZs), we established the binding site of the PBZ molecules using cryo-EM and found that GroEL was the cellular target responsible for the mode of action. Cryo-EM revealed that PBZ1587 binds at the GroEL ring-ring interface (RRI).

The kinase Rio1 and a ribosome collision-dependent decay pathway survey the integrity of 18S rRNA cleavage.

The 18S rRNA sequence is highly conserved, particularly at its 3'-end, which is formed by the endonuclease Nob1. How Nob1 identifies its target sequence is not known, and in vitro experiments have shown Nob1 to be error-prone. Moreover, the sequence around the 3'-end is degenerate with similar sites nearby.

Alexander Friedenstein, Mesenchymal Stem Cells, Shifting Paradigms and Euphemisms.

Six decades ago, Friedenstein and coworkers published a series of seminal papers identifying a cell population in bone marrow with osteogenic potential, now referred to as mesenchymal stem cells (MSCs). This work was also instrumental in establishing the identity of hematopoietic stem cell and the identification of skeletal stem/progenitor cell (SSPC) populations in various skeletal compartments. In recognition of the centenary year of Friedenstein's birth, I review key aspects of his work and discuss the evolving concept of the MSC and its various euphemisms indorsed by changing paradigms in the field.

Enediyne natural product biosynthesis unified by a diiodotetrayne intermediate.

Enediyne natural products are renowned for their potent cytotoxicities but the biosynthesis of their defining 1,5-diyne-3-ene core moiety remains largely enigmatic. Since the discovery of the enediyne polyketide synthase cassette in 2002, genome sequencing has revealed thousands of distinct enediyne biosynthetic gene clusters, each harboring the conserved enediyne polyketide synthase cassette. Here we report that (1) the products of this cassette are an iodoheptaene, a diiodotetrayne and two pentaynes; (2) the diiodotetrayne represents a common biosynthetic intermediate for all known enediynes; and (3) cryptic iodination can be exploited to increase enediyne titers.

Myeloid NCOA4 sequesters KEAP1 to reduce ferroptosis for protection against salmonellosis in mice.

Salmonellosis, caused by serovar Typhimurium, is a significant global threat. Host immunity limits bacterial replication by inducing hepcidin, which degrades ferroportin, reducing iron transfer. However, this boosts macrophage iron storage, aiding intracellular pathogens like .

Synthesis of Membrane-Permeable Macrocyclic Peptides via Imidazopyridinium Grafting.

Macrocyclic peptides (MPs) are a class of compounds that have been shown to be particularly well suited for engaging difficult protein targets. However, their utility is limited by their generally poor cell permeability and bioavailability. Here, we report an efficient solid-phase synthesis of novel MPs by trapping a reversible intramolecular imine linkage with a 2-formyl- or 2-keto-pyridine to create an imidazopyridinium (IP)-linked ring.

Target-based discovery of a broad-spectrum flukicide.

Diseases caused by parasitic flatworms impart a considerable healthcare burden worldwide. Many of these diseases-for example, the parasitic blood fluke infection schistosomiasis-are treated with the drug praziquantel (PZQ). However, PZQ is ineffective against disease caused by liver flukes from the genus Fasciola because of a single amino acid change within the target of PZQ, a transient receptor potential ion channel in the melastatin family (TRPM), in Fasciola species.

Deficiency of the Deubiquitinase UCHL1 Attenuates Pulmonary Arterial Hypertension.

Background: The ubiquitin-proteasome system regulates protein degradation and the development of pulmonary arterial hypertension (PAH), but knowledge about the role of deubiquitinating enzymes in this process is limited. UCHL1 (ubiquitin carboxyl-terminal hydrolase 1), a deubiquitinase, has been shown to reduce AKT1 (AKT serine/threonine kinase 1) degradation, resulting in higher levels. Given that AKT1 is pathological in pulmonary hypertension, we hypothesized that UCHL1 deficiency attenuates PAH development by means of reductions in AKT1.

Heterobifunctional small molecules to modulate RNA function.

RNA has diverse cellular functionality, including regulating gene expression, protein translation, and cellular response to stimuli, due to its intricate structures. Over the past decade, small molecules have been discovered that target functional structures within cellular RNAs and modulate their function. Simple binding, however, is often insufficient, resulting in low or even no biological activity.

Rhes, a striatal enriched protein, regulates post-translational small-ubiquitin-like-modifier (SUMO) modification of nuclear proteins and alters gene expression.

Rhes (Ras homolog enriched in the striatum), a multifunctional protein that regulates striatal functions associated with motor behaviors and neurological diseases, can shuttle from cell to cell via the formation of tunneling-like nanotubes (TNTs). However, the mechanisms by which Rhes mediates diverse functions remain unclear. Rhes is a small GTPase family member which contains a unique C-terminal Small Ubiquitin-like Modifier (SUMO) E3-like domain that promotes SUMO post-translational modification of proteins (SUMOylation) by promoting "cross-SUMOylation" of the SUMO enzyme SUMO E1 (Aos1/Uba2) and SUMO E2 ligase (Ubc-9).

Ironing out the details of ferroptosis.

Ferroptosis, spurred by excess labile iron and lipid peroxidation, is implicated in various diseases. Advances have been made in comprehending the lipid-peroxidation side of ferroptosis, but the exact role of iron in driving ferroptosis remains unknown. Although iron overload is characterized in multiple disease states, the potential role of ferroptosis within them remains undefined.

Mechanisms and consequences of mRNA destabilization during viral infections.

During viral infection there is dynamic interplay between the virus and the host to regulate gene expression. In many cases, the host induces the expression of antiviral genes to combat infection, while the virus uses "host shut-off" systems to better compete for cellular resources and to limit the induction of the host antiviral response. Viral mechanisms for host shut-off involve targeting translation, altering host RNA processing, and/or inducing the degradation of host mRNAs.

microbeMASST: a taxonomically informed mass spectrometry search tool for microbial metabolomics data.

microbeMASST, a taxonomically informed mass spectrometry (MS) search tool, tackles limited microbial metabolite annotation in untargeted metabolomics experiments. Leveraging a curated database of >60,000 microbial monocultures, users can search known and unknown MS/MS spectra and link them to their respective microbial producers via MS/MS fragmentation patterns. Identification of microbe-derived metabolites and relative producers without a priori knowledge will vastly enhance the understanding of microorganisms' role in ecology and human health.

Neuronal knockdown of Cullin3 as a Drosophila model of autism spectrum disorder.

Mutations in Cullin-3 (Cul3), a conserved gene encoding a ubiquitin ligase, are strongly associated with autism spectrum disorder (ASD). Here, we characterize ASD-related pathologies caused by neuron-specific Cul3 knockdown in Drosophila. We confirmed that neuronal Cul3 knockdown causes short sleep, paralleling sleep disturbances in ASD.