Gang Liu

A central dogma of molecular biology is the “speed-accuracy trade-off,” where ribosomes must slow down to ensure accurate protein synthesis. In mycobacteria, a high basal level of mistranslation at glutamine and asparagine codons, caused by an indirect tRNA aminoacylation pathway, promotes tolerance to the antibiotic rifampicin. While pharmacologically increasing translational fidelity is a promising strategy to combat antibiotic tolerance, the underlying mechanisms remain poorly understood. Here, we screened 9,000 synthetic compounds and identified benzo[d]isoxazole-4,7-diones as a novel chemical class that reduces mycobacterial mistranslation. Medicinal chemistry optimization yielded a lead compound, 9787, with superior potency in decreasing mistranslation and reversing rifampicin tolerance. Using competitive chemical proteomics, we identified the 30S ribosomal protein S5 (RpS5) as the specific cellular target. Remarkably, compound 9787 enhances translational fidelity at concentrations that do not measurably impact the overall rate of protein synthesis. Our findings challenge the universality of the speed-accuracy trade-off, demonstrating that fidelity can be improved independently of translation speed. This work reveals that the ribosomal small subunit is a druggable target for modulating translational quality control, and introduces a new strategy for combating antibiotic-tolerant bacteria without the associated fitness cost of slowed translation. IMPORTANCE: A fundamental principle in molecular biology holds that ribosomes face a trade-off between translation speed and accuracy: going faster means making more errors, while maintaining high fidelity requires slowing down. This study challenges that paradigm by identifying a small molecule that increases translational accuracy in mycobacteria without affecting the rate of protein synthesis. The compound targets ribosomal protein S5 and specifically reduces errors arising from physiologically mischarged tRNAs—a quality control problem distinct from the well-studied codon-anticodon mismatches. This form of mistranslation contributes to antibiotic tolerance in tuberculosis, making it a potential therapeutic target. Our findings reveal unexpected flexibility in how ribosomes maintain translation quality and suggest that pharmacologically increasing fidelity without the fitness cost of slowed protein synthesis may be an attractive strategy for combating antibiotic-tolerant bacteria.

Acute pancreatitis (AP) is a sterile inflammatory disease characterized by aberrant zymogen activation, tissue autodigestion, and cross-amplifying inflammatory cascades that complicate targeted therapies. Our single-cell transcriptomics analysis revealed early NOD1 pathway activation in pancreatic acinar and ductal cells in cerulein-induced AP mice, suggesting potential involvement across these cell types. Here, through a scaffold-hopping strategy, we obtained novel-scaffold NOD1 antagonists. Further optimization yielded 32q and its highly water-soluble disodium salt, 32q-2Na. They exhibited low-nanomolar potency against NOD1 with high selectivity. 32q-2Na pharmacologically inhibited NOD1 signaling, significantly reducing serum levels of amylase (AMY), lipase (LIP), MCP-1, and IL-6, and markedly attenuating pancreatic inflammatory responses in cerulein- and FAEE-induced murine models of AP. In conclusion, these findings establish NOD1 as a highly tractable upstream regulatory node for AP intervention.

Despite a century of advances in tuberculosis (TB) diagnostics, a deceptively simple question remains unanswered: Can we reliably detect a single viable Mycobacterium tuberculosis (Mtb) bacillus in a clinical sample? In this perspective, we examine three complementary single-cell measurement platforms that face significant challenges in addressing this seemingly simple yet exceedingly difficult question. Single-cell RNA sequencing (scRNA-seq) can resolve host immune heterogeneity by identifying novel cell subsets and diagnostic biomarkers, such as GBP5, that are otherwise obscured in bulk measurements. Direct Mtb scRNA-seq offers unprecedented insights into four critical biological mechanisms: infectivity, drug resistance, persister formation, and host–pathogen interactions; however, its practical clinical application remains limited. Complementing these genomic approaches, single-cell surface-enhanced Raman spectroscopy serves as a spectroscopic platform to capture molecular heterogeneity among individual bacilli, revealing drug-resistance signatures that are typically masked by population-averaged spectra. Furthermore, single-cell fluorescent probes, arguably the most promising approach for ultimately overcoming this challenge, act as chemical measurement platforms. They enable the direct visualization of viable Mtb by leveraging pathogen-specific metabolic, enzymatic, or transport-based fluorescence activations. We critically evaluate existing translational barriers, including inadequate sensitivity across diverse clinical specimens (e.g., sputum, swabs, body fluids, tissues, and breath aerosols) as well as high costs and incompatibility with routine clinical workflows. Looking ahead, we underscore future direction in TB diagnostic measurement science, emphasizing specimen-driven probe design and the critical challenge of distinguishing slow replication-competent bacilli from metabolically active but nonreplicating populations. Collectively, these emerging technologies chart a roadmap toward the ultimate goal of detecting a single viable Mtb bacterium in clinical samples.

Therapeutic resistance remains a prevalent and intractable clinical challenge across a broad spectrum of human malignancies. Despite extensive investigations, the intricate molecular networks by which the tumor microenvironment (TME) mediates such resistance are not fully understood. In this study, we identified nucleotide-binding oligomerization domain-containing proteins 1 and 2 (NOD1/2) as pivotal regulators of adaptive resistance to diverse antitumor therapies, including immune checkpoint blockade (ICB), adoptive T-cell therapy, and cytotoxic chemotherapy. In murine tumor models, genetic ablation of NOD1/2 or receptor-interacting protein kinase 2 (RIPK2), as well as pharmacological inhibition of RIPK2, remodeled the TME by decreasing immunosuppressive macrophages and boosting CD8⁺ T cell infiltration and cytotoxicity. Mechanistically, NOD1/2 activation in macrophages upregulated programmed death-ligand 1 (PD-L1) expression via the RIPK2/NF-κB signaling axis, establishing an immunosuppressive TME that impaired CD8⁺ T cell-mediated antitumor immunity. Notably, in the clinically relevant setting of immunotherapy resistance, targeted suppression of NOD1/2 signaling in patient-derived peripheral blood mononuclear cells (PBMCs) restored and potentiated ICB responsiveness in patient-derived tumor organoids. Bioinformatic analyses further demonstrated that NOD1/2-associated gene signatures were significantly enriched in tumor-associated macrophages post-therapy. Our findings define NOD1/2 as a novel innate immune checkpoint that orchestrates therapy-induced adaptive resistance and highlight this pathway as a promising target to overcome treatment resistance in refractory cancers.

Emerging SARS‑CoV‑2 variants and related zoonotic sarbecoviruses rely on ACE2 for cell entry, motivating host‑directed antivirals that block spike-ACE2 interaction. Here, we characterize MB‑32, a benzothiazole small molecule that binds ACE2, selectively disrupts binding of SARS‑CoV‑2 spike receptor‑binding domain to ACE2, and preserves ACE2 enzymatic activity across species. MB‑32 potently inhibits entry of SARS‑CoV‑2 variants, SARS‑CoV‑1 and diverse bat/pangolin sarbecoviruses in ACE2‑expressing cells, while sparing vesicular stomatitis virus and authentic MERS‑CoV, indicating non‑virucidal, ACE2‑focused activity. Biochemical and biophysical analyses, supported by ACE2 mutagenesis, support a model in which MB‑32 engages a non‑catalytic surface pocket on the ACE2 N‑terminal helix to allosterically disrupt spike attachment. Intranasal MB‑32 achieves high airway concentrations, protects male ACE2‑transgenic mice and hamsters from SARS‑CoV‑2 disease, and prevents contact transmission of Omicron‑lineage viruses without detectable cardiovascular toxicity. These findings establish MB‑32 as a host‑targeted ACE2 entry inhibitor and provide a framework for small‑molecule ACE2‑directed antivirals against current and future sarbecovirus spillovers.

Microorganisms require iron acquisition, particularly in the form of Fe³⁺ ions from the external environment, for survival. Siderophores are low-molecular-weight compounds secreted by microorganisms that chelate Fe³⁺ ions and facilitate their transport into cells. This process forms an Fe³⁺ uptake system in conjunction with specific transporters, enabling microorganisms to meet their iron requirements. A promising strategy involves utilizing the Fe³⁺ transport mechanism to deliver siderophore-cargo conjugates into microbial cells, thereby enabling the treatment and diagnosis of infectious diseases, including those caused by drug-resistant bacteria. In this perspective, we systematically evaluate each type of siderophore-antibiotic/fluorophore conjugate and its associated therapeutic and diagnostic outcomes. Through this perspective, we aim to further develop the iron uptake strategy as a means to address the increasingly severe global public health challenge of antimicrobial resistance and to achieve rapid and ultrasensitive diagnosis of infections, encompassing but not limited to bacterial and fungal infections.

The repair of DNA double strand break (DSB) is crucial for genome stability. Homologous recombination (HR) is a high-fidelity, error-free DSB repair pathway, and its dysfunction leads to genome instability and can result in tumorigenesis. In HR deficient contexts, microhomology-mediated end joining (MMEJ), dependent on DNA polymerase theta (Polθ), is up-regulated to serve as a backup pathway for DSB repair. Several studies have proved that Polθ inhibition causes synthetic lethality with HR deficiency, and Polθ emerges as a potential DNA damage repair (DDR) target for the treatment of HR deficient tumors both as single agent or in combination with PARP inhibitors (PARPi). In addition, evidences show that the blockage of Polθ can enhance the sensitivity to DNA damage agents. Herein, we characterized ZMS-6659, a highly potent small-molecule Polθ inhibitor targeting the helicase domain. ZMS-6659 induces synthetic lethality in HR deficient models, elicits synergistic effects with even low dose of PARPi and delays PARPi resistance without additional hematotoxicity. Besides, ZMS-6659 also enhance the sensitivity to topoisomerase inhibitors in HR proficient cell lines. These findings support the clinical exploration of ZMS-6659 in combination with PARPi in HR deficient tumors, and suggest a potential application expansion beyond HR deficiency.

Mycobacterium tuberculosis (Mtb) utilizes a heterodimeric ABC transporter (IrtAB) to extract Fe³⁺ ions from host cells. This study demonstrates that ultrasensitive fluorescent probes, achieved through the conjugation of fluorophores with the ligand of IrtAB, N14G and N14G-Fe can detect Mycobacterium smegmatis at concentrations as low as 1-10 nM within an incubation period of less than 5 min. Furthermore, these probes effectively label Mycobacterium bovis Bacille Calmette-Guérin BCG and the wild-type Mtb strain H37Rv at a concentration of 0.1 μM after 10 min of incubation, achieving a limit of detection of 34 Colony-Forming Unit for the wild-type Mtb strain H37Rv. Both N14G and N14G-Fe successfully identified Mtb in sputum samples from patients diagnosed with tuberculosis, exhibiting exceptional fluorescence.

Helicobacter pylori (H. pylori) infection is characterized by the complex interplay between H. pylori and gastric disorders. It has been established that NOD1 can be activated by the peptidoglycan (PGN) present in the cell wall of H. pylori, serving as a key mediator of inflammation and initiating the RIP2/NF-κB and MAPK inflammatory signaling pathways. In this article, we reported on the development of a 2-chloroquinazolin-4-ol derivative 66 as a potent and selective antagonist of both human and mouse NOD1, which effectively inhibited the expression of inflammatory cytokines (IL-6, TNF-α) and chemokines (CXCL1, CXCL8) in immune and epithelial cells, as well as inflammatory cytokines (KC, IL-6) in a H. pylori-induced murine model of gastritis following oral administration. This study laid a foundation for treating gastritis induced by H. pylori infection.

A central dogma of molecular biology is the "speed-accuracy trade-off," where ribosomes must slow down to ensure accurate protein synthesis. In mycobacteria, a high basal level of mistranslation at glutamine and asparagine codons, caused by an indirect tRNA aminoacylation pathway, promotes tolerance to the antibiotic rifampicin. While pharmacologically increasing translational fidelity is a promising strategy to combat antibiotic tolerance, the underlying mechanisms remain poorly understood. Here, we screened 9,000 synthetic compounds and identified benzo[d]isoxazole-4,7-diones as a novel chemical class that reduces mycobacterial mistranslation. Medicinal chemistry optimization yielded a lead compound, 9787 , with superior potency in decreasing mistranslation and reversing rifampicin tolerance. Using competitive chemical proteomics, we identified the 30S ribosomal protein S5 (RpS5) as the specific cellular target. Remarkably, compound 9787 enhances translational fidelity at concentrations that do not measurably impact the overall rate of protein synthesis. Our findings challenge the universality of the speed-accuracy trade-off, demonstrating that fidelity can be improved independently of translation speed. This work reveals that the ribosomal small subunit is a druggable target for modulating translational quality control and introduces a new strategy for combating antibiotic-tolerant bacteria without the associated fitness cost of slowed translation. Importance: A fundamental principle in molecular biology holds that ribosomes face a trade-off between translation speed and accuracy: going faster means making more errors, while maintaining high fidelity requires slowing down. This study challenges that paradigm by identifying a small molecule that increases translational accuracy in mycobacteria without affecting the rate of protein synthesis. The compound targets ribosomal protein S5 and specifically reduce errors arising from physiologically mischarged tRNAs – a quality control problem distinct from the well-studied codon•anticodon mismatches. This form of mistranslation contributes to antibiotic tolerance in tuberculosis, making it a potential therapeutic target. Our findings reveal unexpected flexibility in how ribosomes maintain translation quality and suggest that pharmacologically increasing fidelity without the fitness cost of slowed protein synthesis may be an attractive strategy for combating antibiotic-tolerant bacteria.