Browse Articles

Versatile NTP recognition and domain fusions expand the functional repertoire of the ParB-CTPase fold beyond chromosome segregation

Nucleotide triphosphate (NTP)-dependent molecular switches regulate essential cellular processes by cycling between active and inactive states through nucleotide binding and hydrolysis. These mechanisms were long thought to rely exclusively on ATPase or GTPase proteins, until the discovery of CTPase activity in the bacterial chromosome segregation protein ParB. In the ParAB S system, CTP binding enables ParBs’ accumulation around the centromere-like parS DNA sites to activate the ATPase ParA, thereby facilitating chromosome partitioning to daughter cells. CTP hydrolysis then releases ParB from DNA for recycling. This discovery uncovered a new regulatory principle, but the broader diversity of proteins employing a CTPase mechanism remains unclear. Here, we conduct a large-scale survey of proteins harboring the ParB-CTPase fold across bacteria, archaea, bacteriophages, and eukaryotes. While many ParB-like proteins follow the canonical ParAB S organization with ParA partners, we also identify numerous orphan homologs encoded outside of the parAB operon, frequently linked to mobile genetic elements that may have driven their rapid diversification. The ParB-CTPase folds in these divergent proteins are often fused to lineage-specific domains with diverse predicted biological activities. We further demonstrate that while many homologs retain CTP-binding, others instead bind ATP or GTP, revealing a broader spectrum of nucleotide specificities than previously appreciated. Our findings establish the ParB-CTPase fold as a widely distributed and evolutionarily versatile NTP-binding module, repeatedly co-opted through domain fusion and shifts in nucleotide specificity to enable functions far beyond the classical ParAB S -mediated DNA segregation.

Deciphering guanidine assimilation and riboswitch-based gene regulation in cyanobacteria for synthetic biology applications

Guanidine is well known as a denaturing agent. However, recent studies have demonstrated both the widespread synthesis of guanidine, e.g., in plants and mammals, as well as the widespread occurrence of guanidine metabolism in bacteria, suggesting a broader biological role. Here, we provide insights into guanidine assimilation via guanidine hydrolases (GdmH) in cyanobacteria. The gdmH gene is widespread among cyanobacteria and enables growth on guanidine as the sole nitrogen source. Consistent with this, gdmH gene expression increased under nitrogen limitation, regulated by the transcription factor NtcA. However, guanidine is toxic above 5 mM, necessitating GdmH activity and adaptive mutations activating the multidrug efflux system PrqA. The gdmH gene is frequently colocalized with ABC transporter genes (named gimABC ), which are driven by an additional NtcA-regulated promoter. The corresponding substrate-binding protein GimA showed high affinity to guanidine. Consistent with a high affinity import system, disruption of genes gimA or gimB impaired guanidine-dependent growth of Synechocystis sp. PCC 6803 at low concentrations. However, in presence of >1 mM guanidine, these mutants grew like wildtype, suggesting the existence of additional uptake mechanisms for guanidine. We also demonstrate the high-affinity binding of guanidine to a previously described, conserved RNA motif located within the gdmH 5’-untranslated region, validating it as a guanidine-I riboswitch. By combining it with various promoters, we achieved precise, titratable control of heterologous gene expression in cyanobacteria in vivo. Our findings establish guanidine assimilation as an integral element of cyanobacterial nitrogen metabolism and highlight guanidine riboswitches as valuable tools for synthetic biology.

Structural modeling reveals the allosteric switch controlling the chitin utilization program of <i> <i>Vibrio cholerae</i> </i>

Signal transduction by histidine kinases (HKs) is nearly ubiquitous in bacterial species. HKs can either sense ligands directly or indirectly via a cognate solute-binding protein (SBP). The molecular basis for SBP-dependent signal reception, however, remains poorly understood in most cases. CBP and ChiS are the SBP–HK pair that activate the chitin utilization program of Vibrio cholerae . Here, we elucidate the molecular basis for allosteric regulation of CBP–ChiS by generating structural models of this complex in the unliganded and liganded states, which we support with extensive genetic, biochemical, and cell biological analysis. Our results reveal that ligand-binding induces a large conformational interface switch that is distinct from previously described SBP–HKs. Structural modeling suggests that similar interface switches may also regulate other uncharacterized SBP–HKs. Together, these results extend our understanding of signal transduction in bacterial species and highlight an approach for uncovering the molecular basis of allostery in protein complexes.

Distinct transcription factor interactions drive HOXB13 activity in different stages of prostate cancer

HOXB13 is a lineage-specific transcription factor that plays a critical role in initiation and progression of prostate cancer (PCa). While most research has focused on the role of HOXB13 on androgen receptor (AR) activity, here we demonstrate that HOXB13 is frequently expressed in AR-negative tumors and is essential for the proliferation of both AR-positive and -negative PCa models. Strikingly, HOXB13 is remarkably selective and has almost no effect on nonprostatic tissues. Despite this common essentiality in PCa, HOXB13 activity is markedly different in AR-negative stem cell–like tumors, where interactions with the AP-1 change the HOXB13 cistrome and interactome. Yet despite these distinct activities, HOXB13 activity is commonly mediated by SMARCD2, a member of the mSWI/SNF chromatin remodeling complex. The HOXB13/SMARCD2 interaction alters chromatin accessibility at HOXB13-binding sites, causing increased proliferation in AR-negative PCa. Overall, this work demonstrates a distinct mechanism of action for HOXB13 and highlights its critical role in AR-negative castration-resistant PCa.

A ferritin-like diiron oxygenase BioE initiates bacterial biotin synthesis, a promising antivirulence target

Biotin is an essential enzyme cofactor for intermediary metabolism, and its importance is reflected by the multiplicity of bacterial pathways to its universal precursor, pimelic acid. Here, we report identification of a fourth pimeloyl pathway in the rare but clinically important pathogens Elizabethkingia and Chryseobacterium . This pathway is encoded by two associated structural genes, bioE and bioL . BioE is a ferritin-like nonheme diiron oxygenase that oxidatively cleaves saturated C n (n = 14, 16, 18) fatty acyl coenzyme A (CoA) or acyl carrier protein (ACP) substrates to pimeloyl-CoA/ACP and the free C n-7 acid. The catalytic activity was demonstrated by both in vitro enzymatic assays and the capacity of the bioE gene to complement the genetic defect of an Escherichia coli biotin indicator strain that cannot produce the pimeloyl precursor. BioL, an unusual MocR-type bifunctional transcription factor, negatively regulates bioE expression in response to binding of the downstream intermediate 7-keto-8-aminopelargonic acid. Disruption of bioE in Elizabethkingia meningoseptica and Chryseobacterium indologenes makes them auxotrophic for biotin, impairs biofilm formation, and attenuates bacterial infectivity. Taken together, our findings expand enzymatic diversity of biotin biosynthesis and suggest that selective inhibition of this BioE pathway could provide a therapeutic strategy against recalcitrant nosocomial infections caused by these multidrug-resistant pathogens.

The prion-like characteristic of ORF3 contributes to virion release and pathogenesis of hepatitis E virus

Hepatitis E virus (HEV), the causative agent of hepatitis E, is threatening public health globally. Due to the shortage of efficient in vitro cell culture systems and in vivo model, the viral replication and pathogenesis mechanisms remain largely unknown. Here, we found that HEV-ORF3 protein showed prion-like properties in HEV-infected cells and existed as both monomer and SDS-resistant aggregate forms. In an in vitro cell-free model, incubation of ORF3 monomer with its aggregates could effectively convert ORF3 monomer into aggregates, mirroring a typical characteristic of prion. In addition, the prion domain (PrD) of a classic yeast prion Sup35 could be functionally replaced by full-length HEV-ORF3 or its N-terminal candidate PrD (cPrD). An F10S substitution in the ORF3-cPrD impaired HEV-ORF3 aggregation propensity and blocked the function of ORF3 in enhancing the stability of microtubules in HEV-infected cells, thus led to the inhibition of viral capsid translocation to microtubules and virion release from infected cells. In Mongolian gerbil models, HEV bearing ORF3F10S mutation demonstrated attenuated virulence in vivo compared with wild-type HEV, as evidenced by reduced viremia and viral shedding, as well as alleviated pathological changes of liver tissue in gerbils infected by HEV-ORF3F10S mutant. In conclusion, our data suggest that HEV-ORF3 is a prion-like protein which is involved in viral capsid translocation and virion release, supporting the hypothesis that the self-propagating properties of prion proteins or prion-like proteins are widely exploited in nature and play diverse roles in physiological function.

Prostaglandin E ₂ -EP2/EP4 signaling induces the tumor-infiltrating Treg phenotype for tumor growth

Foxp3 + regulatory T cells (Tregs) heavily infiltrate malignant tumors and restrict antitumor immunity. These tumor-infiltrating Tregs (TI-Tregs) adopt a distinct phenotype by expressing a unique set of genes. This TI-Treg gene expression signature is conserved in TI-Tregs across species and tumor types and stages, suggesting the presence of a common inducing mechanism in the tumor microenvironment (TME). However, identity of such a mechanism remains elusive. Here, we show that prostaglandin E 2 (PGE 2 ) produced in TME directly acts on its receptor EP2/EP4 on Tregs to induce the TI-Treg phenotype. PGE 2 added to TCR-activated Tregs induces a set of genes, many of which are included in the TI-Treg signature, in both induced Tregs (iTregs) and naturally occurring Tregs (nTregs) via EP2/EP4- cAMP-PKA pathway. Concomitantly, PGE 2 -treated Tregs exhibit potent suppressive activity to CD8 + T cells and strongly inhibit their proliferation in an EP4 dependent manner. Consistently, selective loss of EP2 and EP4 in mouse Tregs reduces expression of those genes in Tregs infiltrating Lewis lung carcinoma 1 (LLC1) mouse tumor and significantly delays the tumor progression. In human FOXP3 + iTregs, PGE 2 -EP4 signaling upregulated the expression of Treg signature genes, FOXP3, CD25, and CTLA-4 as well as a typical TI-Treg signature gene, 4-1BB, and enhanced suppressive activity. Furthermore, analysis of single-cell RNA sequencing of nasopharyngeal cancer patients demonstrates preferential expression of the TI-Treg signature genes in Tregs infiltrating the PTGS2 hi tumor group compared to the PTGS2 lo tumor group. These findings suggest that PGE 2 -EP2/EP4 signaling is one of the core mechanisms inducing the TI-Treg phenotype in TME for tumor growth.

LHPP expression in triple-negative breast cancer promotes tumor growth and metastasis by modulating the tumor microenvironment

Triple-negative breast cancer (TNBC) is a highly aggressive and metastatic form of breast cancer that lacks an effective targeted therapy. To identify potential therapeutic targets, we investigated the phosphohistidine phosphatase, LHPP, which has been implicated in the development of several types of cancer. However, the full significance of LHPP in cancer progression remains unclear due to our limited understanding of its molecular mechanism. We found that levels of the LHPP phosphohistidine phosphatase were significantly increased in human breast cancer patients compared to normal adjacent tissues, with the highest levels in the TNBC subtype. When LHPP was knocked out in the MDA-MB-231 human TNBC cell line, cell proliferation, wound healing capacity, and invasion were significantly reduced. However, LHPP knockout in TNBC cells did not significantly affect overall phosphohistidine protein levels. Interestingly, LHPP knockout in MDA-MB-231 cells delayed tumor growth and reduced metastasis when orthotopically transplanted into mouse mammary glands. To investigate LHPP’s role in breast cancer progression, we used next-generation sequencing and proximity-labeling proteomics, and found that LHPP regulates gene expression in chemokine-mediated signaling and actin cytoskeleton organization. Depletion of LHPP reduced the presence of tumor-infiltrating macrophages in mouse xenografts. Our results support a tumor promoter role for LHPP phosphohistidine phosphatase in MDA-MB-231TNBC cells and suggest that targeting LHPP phosphatase could be a potential therapeutic strategy for TNBC.

Absence of higher than sixfold coordination in glassy GeO ₂ up to 158 GPa revealed by X-ray absorption spectroscopy

Simple binary oxide glasses can exhibit a compression behavior distinct from that of their crystalline counterparts. In this study, we employed high-pressure X-ray absorption spectroscopy, coupled to the diamond anvil cell, to investigate in detail local structural changes around Ge in glassy GeO 2 up to 158 GPa. We conducted four independent runs, both with and without pressure-transmitting media. Up to 30 GPa, we observed no significant influence of the pressure medium on the pressure dependence of the Ge–O bond length ( &lt;R Ge–O &gt; ). Between 10 and 30 GPa, the evolution of &lt;R Ge–O &gt; shows substantial variability across our experiments and previous works. The measured values lie close to those reported for crystalline polymorphs, including the rutile- and CaCl 2 -type phase of GeO 2 . This finding suggests that the amorphous structure possesses considerable flexibility to transition among different atomic configurations. From 30 GPa to 158 GPa, our results for both &lt;R Ge–O &gt; and the nonbonded cation–cation distance &lt;R Ge…Ge &gt; demonstrate that edge-sharing octahedra remain the main structural motifs in glassy GeO 2 . Up to 100 GPa, compaction proceeds primarily via distortions of octahedral O–Ge–O bond angles, accompanied by octahedral bond shortening and symmetrization. Above 100 GPa, octahedral distortion becomes the prevailing mechanism. Compared to its crystalline analogues (α-PbO 2 and pyrite-like phase), glassy GeO 2 exhibits a slightly less efficient compaction mechanism, likely due to kinetic constraints that inhibit reconstructive lattice rearrangements.

Data-driven enhanced sampling of mechanistic pathways

The mechanisms of molecular processes can be characterized by following the minimum free energy pathway (MFEP) on the underlying multidimensional conformational landscapes. Despite recent advancements in enhanced sampling algorithms, obtaining a converged high-dimensional molecular free energy landscape remains a considerable challenge. To circumvent this issue, we employ a deep multitask learning algorithm that integrates deep neural networks with the established enhanced sampling method of well-tempered metadynamics to iteratively learn the MFEP between reactant and product conformations, without the knowledge of the underlying free energy landscape. Our approach improves upon existing pathway exploration algorithms by following a simpler protocol, thereby eliminating the need to identify intermediate structures along a guess path. From the learned pathway, an automatic reconstruction of a mechanistic fingerprint can be performed by following the sequence of events in the molecular process, allowing for a direct characterization of the molecular mechanism. We demonstrate applications of our algorithm to prototypical chemical reactions, protein folding, and ligand–receptor binding problems. Due to its low computational cost and overall simplicity, this framework is expected to find widespread applications in elucidating molecular mechanisms at all-atom resolution.

Distinguishing subtypes of endothelial cells in the mouse aorta

This study reanalyzed endothelial cell (EC) gene expression in the mouse aorta utilizing eight single-cell RNA-sequencing datasets. Contrary to the assumption that all ECs originated from the luminal surface, we identified two distinct sites of origin: aortic lumen ( Cytl1 -positive ECs) and peri-aortic microvascular ( Gpihbp1 -positive ECs). The proportions of these EC subtypes varied extensively across the eight datasets, likely due to differing experimental procedures. We deduced complete transcriptomes for each EC subtype, revealing differential gene expression and predicted functional properties. We provide marker lists and an accessible online database for gene-by-gene analysis to aid in correct cell identification and in gauging the impact of patho/physiological parameters on aortic EC gene expression.

Asparagine endopeptidase prompts breast cancer–related pericardial calcification by regulating IGF2 and integrin αvβ5

Cardiac calcification, often seen in age-related diseases, impairs heart function, yet its association with malignant tumors remains largely overlooked. Our study revealed that pericardial calcification (PC) occurs in up to 80% of breast cancer patients with pulmonary metastasis. We demonstrate a reciprocal relationship where breast cancer drives PC, which in turn accelerates cancer progression in humans and mice. Lung metastases increase monocyte-derived macrophage and mesenchymal stem cell (MSC)-derived osteoblast infiltration in the pericardial tissue, triggering inflammation and calcification. Mechanistically, metastatic cancer cells in the lungs highly express and secrete asparagine endopeptidase (AEP), which cleaves IGF2BP3 to free IGF2. AEP and IGF2 contribute to PC by promoting osteoblast differentiation in heart tissue through integrin αvβ5 and IGF1R activation, respectively. Pharmacological blockade of integrin αvβ5 and IGF1R, especially when combined, effectively inhibits ectopic osteogenesis and disrupts the feedback loop between PC and cancer progression. These findings elucidate the interplay between metastatic breast cancer and PC and suggest therapeutic strategies to hinder breast cancer progression.

Carbonate burial regimes, the Meso-Cenozoic climate, and nannoplankton expansion

The long-term climate of Earth alternates between warmer and cooler periods, for which atmospheric CO 2 content is often viewed as a primary control. Although silicate weathering feedback governs this long-term equilibrium, the partitioning of carbonate burial between neritic and pelagic environments can influence how rapidly the carbon cycle adjusts to perturbations. The impact of neritic carbonates accumulation on oceanic calcium and alkalinity fluxes, nannoplankton productivity, and carbon reservoirs is overlooked. To investigate this role, we combine plate-tectonic reconstructions with climatic and physiographic simulations and apply a macroecological model to estimate Meso-Cenozoic neritic warm-water carbonate habitats and productivity. We find that only during the Early Cretaceous and Cenozoic did geochemical influxes exceed the carbonate accumulation potential of neritic platforms, while for most of the interval they fell short. These alternating states define two regimes: i) habitability-limited periods, where environmental conditions restrict the development of neritic carbonate factories, and ii) alkalinity-limited periods, where ocean chemistry restricts carbonate precipitation. By alternating between these regimes, neritic platforms modulate the buffering capacity and the timescales (10 3 to 10 5 years) of carbon cycle recovery and regulate the development and productivity of nannoplankton, and therefore the biological pump. This framework highlights the role of shallow-water carbonate systems as important modulators of the Earth’s climate system and pelagic ecosystems.

Origin and adaptive evolutionary trajectory of the 3′ UTR–derived sRNA UhpU in Enterobacteriaceae

Bacterial small RNAs (sRNAs) derived from mRNA 3′ untranslated regions (3′ UTRs) have emerged as important regulators of gene expression, yet their evolutionary origins and functional diversification remain poorly understood compared to the protein-coding sequences of the same transcripts. In this study, we present a comparative analysis of the biogenesis and regulatory functions of UhpU, a 3′ UTR–derived sRNA from the hexose phosphate transporter gene uhpT , in Escherichia and Salmonella . We show that UhpU likely originated as a processed sRNA generated by RNase E cleavage in Enterobacteriaceae, enabling repression of genes in the hexose phosphotransferase system, thereby contributing to hexose phosphate homeostasis in coordination with its parental gene. In Escherichia , UhpU subsequently evolved the ability to repress mprA , a transcriptional repressor, via a UhpU-binding site introduced by a horizontally acquired DNA fragment that extended the mprA 5′ UTR. After divergence from the most recent common ancestor with Escherichia albertii , the lineage comprising Escherichia coli , Escherichia fergusonii , and Shigella acquired a FliA-dependent promoter within the uhpT coding region, allowing independent transcription of UhpU and establishing it as a dual-biogenesis sRNA. Together, our results outline a stepwise trajectory in which UhpU evolved from a processing-derived metabolic regulator to an sRNA with expanded regulatory connections and a lineage-specific FliA-dependent transcriptional program in Escherichia .

An IMPDH2 variant associated with neurodevelopmental disorder disrupts purine biosynthesis and somite organization

IMP dehydrogenase (IMPDH) controls a key regulatory node in purine biosynthesis. Gain-of-function mutations in human IMPDH2 are associated with neurodevelopmental disorders and neuromuscular symptoms including dystonia, but the developmental mechanisms underlying these defects are unknown. We previously showed that these mutants are insensitive to GTP inhibition and hypothesized that their hyperactivity would affect nucleotide metabolism in vivo. Here, we characterize the metabolic and developmental consequences of the neurodevelopmental disorder-associated IMPDH2 mutant, S160del, in Xenopus tropicalis . We show that expressing S160del but not WT human IMPDH2 disrupts purine pools and somite organization in the developing tadpole. We also show that S160del disrupts in vivo IMPDH filament assembly, a well-described IMPDH regulatory mechanism. Cryo-EM structures show that S160del disrupts filament assembly by destabilizing the dimerization of regulatory Bateman domains. Dimerization of Bateman domains and subsequent filament formation can be restored with a high affinity ligand, but this does not restore sensitivity to GTP inhibition, suggesting S160del also disrupts allostery of IMPDH2 filaments. This work demonstrates that the structural effects of patient IMPDH2 variants can cause disruptions both to nucleotide levels and to the normal development of sensorimotor structures, helping us better understand the physiological basis of disease in these patients.

Widespread promiscuous alkaline phosphatases underscore ancient microbial phosphite utilization

Phosphate is often a limiting resource, directly affecting the availability of key biomolecules such as nucleotides. To cope with phosphate scarcity, bacteria have evolved enzymes that utilize alternative phosphorus compounds, including phosphite (Pt). Although a few enzymes oxidize Pt to produce phosphate, the enzymes responsible for Pt oxidation in many environmental bacteria remain unidentified, and the role of microbial Pt oxidation in the global phosphorus cycle is not yet fully understood. In this study, we performed bioinformatic analyses of three Pt-oxidizing enzymes: the native Pt oxidase, phosphite dehydrogenase (PtxD), and two promiscuous Pt oxidases, alkaline phosphatase (PhoA) and carbon–phosphorus lyase. Among these, PhoA was found to be widely distributed across bacteria since the early stages of their evolution. In contrast, PtxD emerged later in a limited number of bacterial lineages that had lost PhoA. Our biochemical characterizations revealed that most extant and reconstructed ancestral PhoAs tested exhibited Pt oxidation activity. Moreover, disruption of active-site residues diminished Pt oxidase activity in PhoA, while only partially affecting its native function. This promiscuous function of PhoA reveals an overlooked mechanism in bacterial phosphate metabolism and underscores the role of Pt in the cycling of bioavailable phosphorus in ecosystems.

Instantaneous response and quantum geometry of insulators

We present the time-dependent Quantum Geometric Tensor (tQGT) as a comprehensive tool for capturing the geometric character of insulators observable within linear response. We show that tQGT describes the zero-point motion of bound electrons and acts as a generating function for generalized sum rules of electronic conductivity. It therefore enables a systematic framework for computing the instantaneous response of insulators, including optical mass, orbital angular momentum, and dielectric constant. This construction guarantees a consistent approximation across these quantities upon restricting the number of occupied and unoccupied states in a low-energy description of an infinite quantum system. We outline how quantum geometry can be generated in periodic systems by lattice interference and examine spectral weight transfer from small frequencies to high frequencies by creating geometrically frustrated flat bands.

Digital twins come to the life sciences

Reviewers are better equipped to detect fraud than editors

Numerical calculation finds the revised elastic field of an edge dislocation to be incorrect