Since the emergence of molecular tension sensors, the understanding of mechanical forces has advanced substantially. However, visualizing molecular tension in mammalian tissues has remained challenging owing to the technical constraints of the Förster resonance energy transfer (FRET)-based molecular tension sensors. Here, we develop a molecular tension sensor based on circularly permuted EGFP with an elastic linker, which is inserted into either αActinin or αCatenin and then fused with mCherry at the C-terminus to simultaneously visualize tension and the amount of sensor protein. This single-fluorophore tension indicator enables subtle tension changes to be visualized simply from color tone under superresolution microscopy. We further generate H11 knock-in mice expressing these indicators, revealing a molecular-specific regulation of mechanical load within tissues. Thus, our molecular tension indicators provide a powerful approach for probing the complex and heterogeneous regulation of mechanical forces in vivo mammalian systems. FRET-free molecular tension indicators enable high-resolution visualization of tension in vivo, revealing that the regulation of mechanical forces is unexpectedly complex, molecule-specific and heterogeneous in live mouse organs.

Archaea play essential roles in global biogeochemical cycles, yet their diversity in environmental samples remains understudied. Limited coverage of universal primers towards archaea, their very low abundance relative to bacteria in the studied samples, inefficient cell lysis and the incompleteness of reference databases complicate archaeal 16S rRNA gene sequencing. Furthermore, standardization lacks due to the absence of commercially available mock communities. Here, we developed a mock community composed of mixtures of DNA from six archaeal and four bacterial strains with known genome sizes and 16S rRNA gene copy numbers. Five mock compositions with different archaeal–bacterial abundance ratios were used to evaluate two universal primer pairs and eight archaea-specific methods. The universal primer pairs detected all taxa but consistently overestimated bacterial abundance, indicating to be less suitable for environments where archaea are more dominant. Archaeal specific primer pairs showed variable specificity, with some exhibiting substantial bacterial co-amplification under certain conditions. The Arch349F–A806R primer pair demonstrated the highest archaeal specificity, the least technical variability, the most consistent agreement with expected mock compositions, and minimal artefactual amplification when applied to a complex bacterial mock community. To show its applicability on environmental samples, this method was applied on Boom Clay pore water samples, which indicated a much higher archaeal diversity than previously shown with a standard commercial method. Overall, this study showed the necessity of primer validation through mock communities and identified Arch349F–A806R as a promising primer pair for archaeal 16S rRNA gene sequencing, especially in samples containing methanogenic archaea.

Methane emissions from rice paddies are a major source of greenhouse gases, and silicate fertilizer (SF) derived from blast furnace slag has emerged as an environmentally friendly amendment that reduces methane release. SF modifies the soil redox environment by supplying electron acceptors such as iron, but the microbial mechanisms underlying methane mitigation remain unclear. Here, we combined metagenomic profiling, microbial isolation, and greenhouse experiments to show that SF consistently enriched Burkholderia vietnamiensis together with the methanotrophs in the rice rhizosphere. Inoculation of B. vietnamiensis strain HU01 alone reproduced the methane-suppressing effect of SF and increased methanotroph abundance. Cell-free culture supernatants of HU01 stimulated the growth of Methylococcus geothermalis and Methylocystis parvus through secreted, low-molecular-weight, heat-stable metabolites. Although metabolic modeling predicted primary metabolites such as L-glutamate and cobalamin as candidate cross-feeding factors, LC–MS/MS analysis of the highly active fractions indicated that the robust bioactivity was primarily driven by a chemically complex metabolite mixture rather than these ubiquitous single compounds. These results provide direct evidence that enrichment of a single heterotrophic species can enhance methane oxidation by stimulating methanotroph activity through metabolite exchange. B. vietnamiensis HU01 represents a promising microbial resource for methane mitigation and illustrates how targeted heterotroph–methanotroph interactions can be leveraged for climate-smart rice cultivation.

Episomal plasmids are widely used in yeast metabolic engineering for their flexibility and high transformation efficiency. However, their application in Yarrowia lipolytica is often limited by low copy number and insufficient genetic stability. In this study, a robust episomal plasmid that balanced genetic stability and protein expression capacity was developed in Y. lipolytica. The autonomously replicating sequence (ARS) elements were selected, and ARS1 was identified as a favorable element that balanced genetic stability and gene expression capacity. To improve plasmid copy number, different ubiquitin degrons were used to tune selection marker abundance, and Ubi-R-Δk-mediated marker resulted in an approximately fourfold increase in plasmid copy number (from ∼1 to ∼5 copies per cell). The copy number of the engineered plasmid was one of the highest‑copy episomal systems in Y. lipolytica. To further evaluate the engineered plasmid, we applied it to β‑carotene biosynthesis. The results showed that the β‑carotene titer was 3.43‑fold higher than that of the non‑optimized plasmid. Collectively, this work expands the episomal plasmid toolbox for Y. lipolytica and establishes a generalizable framework for robust protein expression plasmid engineering in non-conventional yeasts.

Nanopore protein sequencing has advanced from a theoretical ambition to a practical technology, overcoming major challenges in protein unfolding, transport and signal detection to achieve single-amino-acid and post-translational modification sensitivity. With both full-length and peptide-based approaches progressing towards commercialization, the field is now focused on scaling throughput and refining analysis tools to unlock a new generation of single-molecule proteomics.

Curiosity-driven studies of lipid organization and lipid asymmetry in biological membranes led to scalable methods for making liposomes, efficient loading of drugs and ionizable cationic lipids. In turn, these advances led to lipid nanoparticles enabling the RNA medicine Onpattro and the COVID-19 mRNA vaccines, and they are now facilitating gene therapies that can potentially treat most human diseases. lnp

Using near-infrared photoswitchable photochromic genetically encoded probes, differential photoacoustic tomography (PAT) enables deep-tissue imaging with high resolution and sensitivity; however, current bacteriophytochrome-based probes are bulky multidomain proteins exceeding 55 kDa, which constrains their use for designing PAT molecular reporters. Here we engineer rsmiRFPnano, a compact cyanobacteriochrome-derived photoswitchable PAT probe. With a molecular weight below 19 kDa, rsmiRFPnano efficiently incorporates biliverdin, exhibits high extinction coefficients in both photochromic states and demonstrates robust performance in live cells. We established rsmiRFPnano-based reporters for NF-κB and AP-1 pathways, which display low basal activity and strong activation-dependent responses. These reporters effectively visualized acute inflammation using noninvasive PAT in vivo, further validated through cellular-level spatial omics mapping. We then created photoswitchable nanobody probes, rsNIR-Fbs, stabilized upon antigen binding and rapidly degraded otherwise, enabling selective photoacoustic immuno-imaging of intracellular antigens in vivo. Together, these results establish rsmiRFPnano as an ultracompact and versatile platform for molecular PAT. A small, photoswitchable and genetically encodable protein operating in the near-infrared window is reported for photoacoustic tomography applications.