Huilin Li

ZNFX1 is an RNA helicase that plays a critical role in host defense against viral infections. Recent studies further revealed that ZNFX1 harbors an unexpected RNA-ubiquitinating E3 ligase activity, raising fundamental questions about how RNA-binding activates this multifunctional enzyme. Here, we combine cryo-electron microscopy with biochemical analyses to define the structural basis of ZNFX1 regulation. In the absence of RNA, ZNFX1 adopts an autoinhibited conformation in which a blocking helix occludes the RNA-binding groove. It exists predominantly as autoinhibited monomers, and, at higher concentrations, assembles into short helical filaments composed of repeating tetramers. Binding of single-stranded RNA (ssRNA) remodels these higher-order assemblies into RNA-bound ZNFX1 monomers and dimers. The cryo-EM structure of the ssRNA-bound ZNFX1 dimer reveals that RNA binding displaces the blocking helix and induces large-scale conformational rearrangements that remodel the helicase core into an active configuration. Biochemical analyses validate the proposed RNA-binding interfaces and demonstrate ATP-dependent RNA unwinding by ZNFX1. Together, these findings support a regulatory mechanism in which autoinhibited ZNFX1 exists as monomers or concentration-dependent higher-order oligomers, whereas RNA binding promotes structural remodeling and helicase activation.

Malaria remains a significant global health threat, affecting millions of lives each year. The causative agents, Plasmodium parasites, are highly resilient and have developed resistance to nearly all existing antimalarial drugs. The Plasmodium 20S proteasome core (Pf20S), a central component of the parasite’s proteostasis pathway, has emerged as a promising therapeutic target. Inhibiting Pf20S effectively suppresses parasite growth at multiple stages of the parasite life cycle, synergizes with artemisinin-based therapies, and shows no cross-resistance with other antimalarial drug classes. Previously, we identified a novel class of proteasome inhibitors with potent activity against Pf20S, which we subsequently optimized for improved potency and selectivity over human proteasomes. However, these inhibitors exhibited suboptimal pharmacokinetic properties, potentially attributable to a high number of rotatable bonds and hydrogen bond donors. Here, we describe further optimization of this inhibitor class and provide structural insights into Pf20S-inhibitor interactions. Cryo-EM structural studies reveal a novel binding pose of the inhibitor at the Pf20S β5 active site, explaining the enhanced activity of an N-cap sulfonamide modification compared to an N-cap amide. These findings offer insights into the development of next-generation antimalarial Pf20S-targeting compounds with improved drug-like properties.

Guanine-rich nucleic acid sequences can fold into G-quadruplex (G4) structures that regulate DNA replication, transcription, and translation. Fanconi anemia group J helicase (FANCJ) resolves G4 structures at stalled replication forks. Despite its central role in genome maintenance, the molecular basis of G4 recognition and unwinding by FANCJ has remained unclear. Here, we report cryo-EM structures of human FANCJ bound to a G4-containing DNA substrate and ATPγS. The structures reveal direct engagement of the G4 by the Fe–S domain. Structure-guided mutagenesis demonstrates that this interface is essential for G4 binding and unwinding. The structures further capture open and closed conformational states linked to ATP hydrolysis, providing a mechanism for directional translocation along 5′ ssDNA and progressive G4 unwinding. Together, these findings establish the structural basis of G4 recognition by FANCJ and provide mechanistic insights into how disease-associated mutations linked to Fanconi anemia and breast cancer impair helicase function.

Two‐component systems (TCS) mediate bacterial signal transduction in response to specific environmental conditions. The two components are the sensor kinase (SK), which senses the signal and autophosphorylates on a histidine residue, and a response regulator (RR), which is phosphorylated by the kinase and modifies gene expression. Despite intensive study, the mechanisms of signal sensing by sensor kinases remain incompletely defined, and it is yet unclear how these kinases sense multiple ligands. Mycobacterium tuberculosis PdtaS/PdtaR is a soluble TCS pair that participates in the Rip1 signal transduction cascade to control virulence by responding to copper and nitric oxide (NO). In contrast to paradigmatic ligand‐activated SKs, PdtaS is constitutively active in the absence of a ligand and is directly inhibited by Cu or NO, yet it remains unclear how such chemically diverse ligands are sensed. Here, we show that PdtaS is a dimeric kinase that constitutively autophosphorylates in trans . Cu and NO both inhibit PdtaS phosphorylation by inhibiting dimerization. Phylogenetic analysis of the PdtaS family reveals conservation of the GAF/PAS dimer interface rather than the ligand‐binding pockets. Mutations in the GAF dimer interface that alter dimerization impair multi‐ligand sensing both in vitro and in M. tuberculosis cells. These results indicate that a single bacterial kinase can sense chemically diverse inputs through inhibition of dimerization‐dependent phosphorylation.

Reply to Roske and Yeeles: Mismatch correction by a replicative polymerase constrained on DNA by a ring

Background: Chronic kidney disease (CKD) has become a significant issue in global public health, with its prevalence showing a persistent upward trend. Traditional risk factors cannot fully explain the persistent increase in incidence rates. Hypothyroidism, a common endocrine disorder, is considered a potential non-traditional risk factor for CKD, but the evidence remains inconsistent. This study aims to systematically evaluate the association between hypothyroidism and CKD risk. Methods: Search PubMed, Embase, Web of Science, and Cochrane Library databases for relevant literature up to September 1, 2025. Included observational studies reporting the association between overt or subclinical hypothyroidism and CKD risk, providing odds ratios (OR) or hazard ratios (HR) with 95% confidence intervals (CIs). Quality assessment was conducted using the Newcastle–Ottawa Scale (NOS) and the Agency for Healthcare Research and Quality (AHRQ) quality assessment tool. Calculate pooled risk estimates using random-effects models, and conduct subgroup analyses, sensitivity analyses, and publication bias analyses. Results: A total of 13 publications reporting 15 studies (11 cross-sectional and 4 cohort studies) involving approximately 5,101,102 participants were included. The pooled analysis of cross-sectional studies demonstrated that hypothyroidism was significantly associated with CKD (OR = 1.66, 95% CI: 1.43–1.94; I ² = 78.6%). Subgroup analyses revealed consistent associations across different thyroid disease types, regions, diagnostic methods, and study quality group. Pooled results from cohort studies also demonstrated that hypothyroidism significantly increased CKD risk (HR = 1.21, 95% CI 1.08-1.36; I ² = 50.08%). Sensitivity analysis results remained stable. Publication bias was present in cross-sectional studies, but the association remained significant after using the trim-and-fill method (adjusted OR = 1.29, 95% CI: 1.06–1.57). Conclusion: This systematic review and meta-analysis suggests that hypothyroidism is associated with an increased risk of CKD. Future well-designed prospective and interventional studies are needed to clarify temporal and causal relationships and to elucidate the underlying mechanisms between hypothyroidism and renal dysfunction. Systematic Review Registration: https://www.crd.york.ac.uk/prospero/ , identifier CRD420251143834.