Science Journals

Cre-loxP technology, a cornerstone in fate mapping and in vivo gene function studies, faces challenges in achieving precise and efficient conditional mutagenesis through inducible systems. This study introduces two innovative genetic tools designed to overcome these limitations. The first, roxCre, enables DreER-mediated Cre release, paving the way for intersectional genetic manipulation that permits increased precision and efficiency. The second, loxCre, facilitates conditional gene targeting by allowing CreER lines to induce Cre expression with significantly enhanced efficiency. These tools incorporate a fluorescent reporter for genetic lineage tracing, simultaneously revealing efficient gene knockout in cells marked by the reporter. These strategies hold great potential for precise and efficient exploration of lineage-specific gene functions, marking a significant advancement in genetic research methodologies.

Tripartite multidrug efflux pumps that span the cell envelope are essential for antibiotic resistance in Gram-negative bacteria. Here, we report cryo-EM structures of two endogenous efflux complexes from <i>Escherichia coli</i>: a TolC–YbjP subcomplex at 3.56 Å resolution and the complete TolC–YbjP–AcrABZ pump at 3.39 Å. Structural analysis reveals that YbjP, a previously uncharacterized lipoprotein, binds TolC in a 3:3 stoichiometry, bridging the TolC protomers at their equatorial domain. Clear density of the mature YbjP’s N-terminal Cys19 indicates that YbjP is anchored to the outer membrane by an N-terminal lipid moiety. Notably, YbjP remains bound as TolC undergoes AcrA-induced opening, suggesting that this accessory protein accommodates the conformational change. The AcrB trimer simultaneously presents three distinct conformational states (L, T, and O), capturing a complete transport cycle. These high-resolution structures provide insights into the architecture and mechanism of clinically relevant efflux machinery, identifying YbjP as a previously unrecognized structural component that contributes to TolC positioning, and may assist in its membrane localization.

mRNA translation involves multiple regulatory steps, but how translation elongation influences protein output remains unclear. Using SunTag live-cell imaging and mathematical modeling, we quantified translation dynamics in single mRNAs across diverse coding sequences. Our Totally Asymmetric Exclusion Process (TASEP)-based Hidden Markov Model revealed a strong coordination between initiation and elongation rates, resulting in consistently low ribosome density (≤12% occupancy) across all reporters. This coupling persisted under pharmacological inhibition of the elongation factor eIF5A, where proportional decreases in both initiation and elongation rates maintained homeostatic ribosome density. In contrast, eIF5A knockout cells exhibited a significant decrease in ribosome density, suggesting altered coordination. Together, these results highlight a dynamical coupling of initiation and elongation rates at the single-mRNA level, preventing ribosome crowding and maintaining translational homeostasis in mammalian cells.

Humans perform tasks involving the manipulation of inputs regardless of how these signals are perceived by the brain, thanks to representations that are invariant to the stimulus modality. In this paper, we present modality-agnostic decoders that leverage such modality-invariant representations to predict which stimulus a subject is seeing, irrespective of the modality in which the stimulus is presented. Training these modality-agnostic decoders is made possible thanks to our new large-scale fMRI dataset SemReps-8K, released publicly along with this paper. It comprises six subjects watching both images and short text descriptions of such images, as well as the conditions during which the subjects were imagining visual scenes. We find that modality-agnostic decoders can perform as well as modality-specific decoders and even outperform them when decoding captions and mental imagery. Furthermore, a searchlight analysis revealed that large areas of the brain contain modality-invariant representations. Such areas are also particularly suitable for decoding visual scenes from the mental imagery condition.

Fluid flow has a major effect on the aggregation and fragmentation of bacterial colonies. Yet, a generic framework to understand and predict how hydrodynamics affects colony size remains elusive. This study investigates how fluid flow affects the formation and maintenance of large colonial structures in cyanobacteria, using an experimental technique that precisely controls hydrodynamic conditions. We performed experiments on laboratory cultures and lake samples of the cyanobacterium <i>Microcystis</i>, while their colony size distribution was measured simultaneously by direct microscopic imaging. We demonstrate that extracellular polymeric substances (EPS)-embedded cells formed by cell division exhibit significant mechanical resistance to shear forces. However, at elevated hydrodynamic stress levels (exceeding those typically generated by surface wind mixing), these colonies experience fragmentation through an erosion process. We also show that single cells can aggregate into small colonies due to fluid flow. However, the structural integrity of these flow-induced colonies is weaker than that of colonies formed by cell division. We provide a mathematical analysis to support the experiments and demonstrate that a population model with two categories of colonies describes the measured size distributions. Our results shed light on the specific conditions wherein flow-induced fragmentation and aggregation of cyanobacteria are decisive and indicate that colony formation under natural conditions is mainly driven by cell division, although flow-induced aggregation could play a role in dense bloom events. These findings can be used to improve prediction models and mitigation strategies for toxic cyanobacterial blooms and also offer potential applications in other areas, such as algal biotechnology or medical settings where the dynamics of biological aggregates play a significant role.

Cone-rod dystrophy (CRD) is a macular degeneration disorder characterized by initial cone cell degeneration. Mutations in CDHR1, a photoreceptor-specific cadherin, have been found to be associated with the incidence of CRD. While studying the function of CDHR1, we observed that the localization of the zebrafish homologue, cdhr1a, resembles that of calyceal process (CPs). When co-labeling CPs using pcdh15b, we observed that cdhr1a, in the outer segment (OS), juxtaposes with pcdh15b, found in the CP. Similar localization patterns were detected in human, macaque, xenopus, ducks, gerbil, and mouse. Using immunoprecipitation and K652 cell aggregation assays, we demonstrate that pcdh15b and cdhr1a can interact and thus potentially link the OS and CP. To analyze the consequences of OS-CP interactions in CRD, we established a <i>cdhr1a</i> mutant line (<i>cdhr1a^fs*146</i>). Homozygous <i>cdhr1a^fs*146</i> mutants exhibit minor cone OS defects starting at 15 dpf and severe OS disruption and cell loss by 3 months. Shortening of CPs coincided with cone OS defects which were significantly exacerbated when combined with the loss of pcdh15b. Rod OS defects were mild and delayed until 3–6 months. In conclusion, we propose that cdhr1a and pcdh15b function to link cone OSs with CPs and maintain OS integrity.

Intermittent fasting (IF) has emerged as a powerful dietary intervention with profound metabolic benefits, yet the tissue-specific molecular mechanisms underlying these effects remain poorly understood. In this study, we employed comprehensive proteomics and transcriptomics analysis to investigate the systemic and organ-specific adaptations to IF in male C57BL/6 mice. Following a 16 hr daily fasting regimen (IF16) over 4 months, IF reduced blood glucose, HbA1c, and cholesterol levels while increasing ketone bodies, indicative of enhanced metabolic flexibility. Proteomic profiling of the liver, skeletal muscle, and cerebral cortex revealed tissue-specific responses, with the liver exhibiting the most pronounced changes, including upregulation of pathways involved in fatty acid oxidation, ketogenesis, and glycan degradation, and downregulation of steroid hormone and cholesterol metabolism. In muscle, IF enhanced pyruvate metabolism, fatty acid biosynthesis, and AMPK signaling, while suppressing oxidative phosphorylation and thermogenesis. The cerebral cortex displayed unique adaptations, with upregulation of autophagy, PPAR signaling, and metabolic pathways, and downregulation of TGF-beta and p53 signaling, suggesting a shift toward energy conservation and stress resilience. Notably, Serpin A1c emerged as the only protein commonly upregulated across all three tissues, highlighting its potential role in systemic adaptation to IF. Integrative transcriptomic and proteomic analyses revealed partial concordance between mRNA and protein expression, underscoring the complexity of post-transcriptional regulation. Shared biological signaling processes were identified across tissues, suggesting unifying mechanisms linking metabolic changes to cellular communication. Our findings reveal both conserved and tissue-specific responses by which IF may optimize energy utilization, enhance metabolic flexibility, and promote cellular resilience.

A third of patients with the pediatric cerebellar tumor Medulloblastoma (MB) have mutations that activate Sonic hedgehog (SHH) signaling (SHH-MB subgroup). The contribution of secondary mutations to tumor severity, however, is not clear. <i>PTEN</i> mutations are enriched in the SHH-1 subtype that has the lowest survival rate. Widespread heterozygous loss of <i>Pten</i> in two SHH-MB mouse models increases penetrance and accelerates onset of differentiated tumors. We delineated cellular and transcriptional changes that accelerate tumor growth and cause differentiation using a sporadic SHH-MB mouse model expressing oncogenic SmoM2 in rare cerebellar granule cell precursors (GCPs) and scRNA-seq analysis. Homozygous but not heterozygous sporadic loss of <i>Pten</i> resulted in rapid acceleration of tumor growth and end-stage disease by 40 days, compared to ~25% survival in control SmoM2 mice at 100 days. Heterozygous <i>PTEN</i> mutations, therefore, should negatively impact disease outcome primarily with germline mutations. Loss of <i>Pten</i> in normal or SmoM2-expressing GCPs increased proliferation and enhanced progenitor state initially, but by 12 days <i>Pten</i> mutant SmoM2 tumors were highly differentiated due to increased survival of non-proliferating GCPs. Furthermore, macrophage infiltration and cytotoxicity appeared reduced in differentiated regions of tumors lacking <i>Pten</i>, indicating cell nonautonomous changes could also contribute to accelerated tumor growth.

The mechanisms guiding nocturnal insect migration remain poorly understood. Although many species are thought to use the geomagnetic field, the sensory basis of magnetic orientation in insects has yet to be clarified. We developed an indoor experimental system to investigate the integration of geomagnetic and visual cues in the seasonal orientation of a globally distributed pest moth, the fall armyworm (<i>Spodoptera frugiperda</i>), a highly invasive species which in the past decade has colonized almost all potentially habitable regions of the globe. Our results demonstrate that fall armyworms require both geomagnetic and visual cues for accurate migratory orientation, with visual cues being indispensable for magnetic orientation. When visual and geomagnetic cues are placed in conflict, moths become disoriented, although not immediately, indicating that sensory recognition of the conflict requires time to process. We also show that the absence of visual cues leads to a significant loss of flight stability, which likely explains the disruption in orientation. Our findings highlight that visual cues are critical for stable magnetic orientation in the fall armyworm, offering a basis for future investigations of visual-magnetic integration in noctuid migrants.

The transcription factor (TF) regulatory network in <i>Pseudomonas aeruginosa</i> is complex and involves multiple regulators that respond to various environmental signals and physiological cues by regulating gene expression. However, the biological functions of at least half of its 373 putative TFs remain uncharacterised. Herein, chromatin immunoprecipitation sequencing (ChIP-seq) was used to investigate the binding sites of 172 TFs in the <i>P. aeruginosa</i> PAO1 strain. The results revealed 81,009 significant binding peaks in the genome, more than half of which were located in the promoter regions. To further decode the diverse regulatory relationships among TFs, a hierarchical network was assembled into three levels: top, middle, and bottom. Thirteen ternary regulatory motifs revealed flexible relationships among TFs in small hubs, and a comprehensive co-association atlas was established, showing the enrichment of seven core associated clusters. Twenty-four TFs were identified as the master regulators of virulence-related pathways. The pan-genome analysis revealed the conservation and evolution of TFs in <i>P. aeruginosa</i> complex and other species. A web-based database combining existing and new data from ChIP-seq and the high-throughput systematic evolution of ligands by exponential enrichment was established for searching TF-binding sites. This study provides important insights into the pathogenic mechanisms of <i>P. aeruginosa</i> and related bacteria and is expected to contribute to the development of effective therapies for infectious diseases caused by this pathogen.

Temperature is a critical factor influencing the outbreak and progression of viral diseases in organisms. Febrile temperatures have been shown to enhance immune competence and reduce viral replication in various species. However, the underlying mechanisms remain largely unknown. In this study, we investigate the molecular mechanisms by which elevated temperatures confer resistance to viral infections, focusing on the role of heat shock factor 1 (HSF1) in regulating antimicrobial effectors rather than the traditional target genes molecular chaperones. Using shrimp <i>Litopenaeus vannamei</i> as a model, we demonstrate that febrile temperatures induce HSF1, which in turn upregulates antimicrobial peptides (AMPs) that target viral envelope proteins and inhibit viral replication. Importantly, this is the first to show that HSF1 directly binds to the heat shock element (HSE) motifs of AMPs both in shrimp and <i>Drosophila</i> S2 cells, suggesting this may be a conserved regulatory mechanism in arthropods. Additionally, our findings highlight the role of HSF1 beyond the classical heat shock response, revealing its critical function in modulating innate immunity. These insights provide new avenues for managing viral infections in aquaculture and other settings by leveraging environmental temperature control.