Base-resolution m5C profiling across the mammalian transcriptome by bisulfite-free enzyme-assisted chemical labeling approach.

5-methylcytosine (m5C) is a prevalent RNA modification crucial for gene expression regulation. However, accurate and sensitive m5C sites identification remains challenging due to severe RNA degradation and reduced sequence complexity during bisulfite sequencing (BS-seq). Here, we report m5C-TAC-seq, a bisulfite-free approach combining TET-assisted m5C-to-f5C oxidation with selective chemical labeling, therefore enabling direct base-resolution m5C detection through pre-enrichment and C-to-T transitions at m5C sites. With m5C-TAC-seq, we comprehensively profiled the m5C methylomes in human and mouse cells, identifying a substantially larger number of confident m5C sites. Through perturbing potential m5C methyltransferases, we deciphered the responsible enzymes for most m5C sites, including the characterization of NSUN5's involvement in mRNA m5C deposition. Additionally, we characterized m5C dynamics during mESC differentiation. Notably, the mild reaction conditions and preservation of nucleotide composition in m5C-TAC-seq allow m5C detection in chromatin-associated RNAs. The accurate and robust m5C-TAC-seq will advance research into m5C methylation functional investigation.

Recognition of methamphetamine and other amines by trace amine receptor TAAR1.

Trace amine-associated receptor 1 (TAAR1), the founding member of a nine-member family of trace amine receptors, is responsible for recognizing a range of biogenic amines in the brain, including the endogenous ß-phenylethylamine (ß-PEA)1 as well as methamphetamine2, an abused substance that has posed a severe threat to human health and society3. Given its unique physiological role in the brain, TAAR1 is also an emerging target for a range of neurological disorders including schizophrenia, depression and drug addiction2,4,5. Here we report structures of human TAAR1-G-protein complexes bound to methamphetamine and ß-PEA as well as complexes bound to RO5256390, a TAAR1-selective agonist, and SEP-363856, a clinical-stage dual agonist for TAAR1 and serotonin receptor 5-HT1AR (refs. 6,7). Together with systematic mutagenesis and functional studies, the structures reveal the molecular basis of methamphetamine recognition and underlying mechanisms of ligand selectivity and polypharmacology between TAAR1 and other monoamine receptors. We identify a lid-like extracellular loop 2 helix/loop structure and a hydrogen-bonding network in the ligand-binding pockets, which may contribute to the ligand recognition in TAAR1. These findings shed light on the ligand recognition mode and activation mechanism for TAAR1 and should guide the development of next-generation therapeutics for drug addiction and various neurological disorders.

Layer Hall effect in a 2D topological axion antiferromagnet.

Whereas ferromagnets have been known and used for millennia, antiferromagnets were only discovered in the 1930s1. At large scale, because of the absence of global magnetization, antiferromagnets may seem to behave like any non-magnetic material. At the microscopic level, however, the opposite alignment of spins forms a rich internal structure. In topological antiferromagnets, this internal structure leads to the possibility that the property known as the Berry phase can acquire distinct spatial textures2,3. Here we study this possibility in an antiferromagnetic axion insulator-even-layered, two-dimensional MnBi2Te4-in which spatial degrees of freedom correspond to different layers. We observe a type of Hall effect-the layer Hall effect-in which electrons from the top and bottom layers spontaneously deflect in opposite directions. Specifically, under zero electric field, even-layered MnBi2Te4 shows no anomalous Hall effect. However, applying an electric field leads to the emergence of a large, layer-polarized anomalous Hall effect of about 0.5e2/h (where e is the electron charge and h is Planck's constant). This layer Hall effect uncovers an unusual layer-locked Berry curvature, which serves to characterize the axion insulator state. Moreover, we find that the layer-locked Berry curvature can be manipulated by the axion field formed from the dot product of the electric and magnetic field vectors. Our results offer new pathways to detect and manipulate the internal spatial structure of fully compensated topological antiferromagnets4-9. The layer-locked Berry curvature represents a first step towards spatial engineering of the Berry phase through effects such as layer-specific moiré potential.

Comprehensive spatiotemporal mapping of single-cell lineages in developing mouse brain by CRISPR-based barcoding.

A fundamental interest in developmental neuroscience lies in the ability to map the complete single-cell lineages within the brain. To this end, we developed a CRISPR editing-based lineage-specific tracing (CREST) method for clonal tracing in Cre mice. We then used two complementary strategies based on CREST to map single-cell lineages in developing mouse ventral midbrain (vMB). By applying snapshotting CREST (snapCREST), we constructed a spatiotemporal lineage landscape of developing vMB and identified six progenitor archetypes that could represent the principal clonal fates of individual vMB progenitors and three distinct clonal lineages in the floor plate that specified glutamatergic, dopaminergic or both neurons. We further created pandaCREST (progenitor and derivative associating CREST) to associate the transcriptomes of progenitor cells in vivo with their differentiation potentials. We identified multiple origins of dopaminergic neurons and demonstrated that a transcriptome-defined progenitor type comprises heterogeneous progenitors, each with distinct clonal fates and molecular signatures. Therefore, the CREST method and strategies allow comprehensive single-cell lineage analysis that could offer new insights into the molecular programs underlying neural specification.

Precision environmental health monitoring by longitudinal exposome and multi-omics profiling.

Conventional environmental health studies have primarily focused on limited environmental stressors at the population level, which lacks the power to dissect the complexity and heterogeneity of individualized environmental exposures. Here, as a pilot case study, we integrated deep-profiled longitudinal personal exposome and internal multi-omics to systematically investigate how the exposome shapes a single individual's phenome. We annotated thousands of chemical and biological components in the personal exposome cloud and found they were significantly correlated with thousands of internal biomolecules, which was further cross-validated using corresponding clinical data. Our results showed that agrochemicals and fungi predominated in the highly diverse and dynamic personal exposome, and the biomolecules and pathways related to the individual's immune system, kidney, and liver were highly associated with the personal external exposome. Overall, this data-driven longitudinal monitoring study shows the potential dynamic interactions between the personal exposome and internal multi-omics, as well as the impact of the exposome on precision health by producing abundant testable hypotheses.

DCLK1 mediated cooperative acceleration of EMT by avian leukosis virus subgroup J and Marek's disease virus via the Wnt/ß-catenin pathway promotes tumor metastasis.

Co-infection with oncogenic retrovirus and herpesvirus significantly facilitates tumor metastasis in human and animals. Co-infection with avian leukosis virus subgroup J (ALV-J) and Marek's disease virus (MDV), which are typical oncogenic retrovirus and herpesvirus, respectively, leads to enhanced oncogenicity and accelerated tumor formation, resulting in increased mortality of affected chickens. Previously, we found that ALV-J and MDV cooperatively promoted tumor metastasis. However, the molecular mechanism remains elusive. Here, we found that doublecortin-like kinase 1 (DCLK1) mediated cooperative acceleration of epithelial-mesenchymal transition (EMT) by ALV-J and MDV promoted tumor metastasis. Mechanistically, DCLK1 induced EMT via activating Wnt/ß-catenin pathway by interacting with ß-catenin, thereby cooperatively promoting tumor metastasis. Initially, we screened and found that DCLK1 was a potential mediator for the cooperative activation of EMT by ALV-J and MDV, and enhanced cell proliferation, migration, and invasion. Subsequently, we revealed that DCLK1 physically interacted with ß-catenin to promote the formation of the ß-catenin-TCF4 complex, inducing transcription of the Wnt target gene, c-Myc, promoting EMT by increasing the expression of N-cadherin, Vimentin, and Snail, and decreasing the expression of E-cadherin. Taken together, we discovered that jointly activated DCLK1 by ALV-J and MDV accelerated cell proliferation, migration and invasion, and ultimately activated EMT, paving the way for tumor metastasis. This study elucidated the molecular mechanism underlying cooperative metastasis induced by co-infection with retrovirus and herpesvirus. IMPORTANCE: Tumor metastasis, a complex phenomenon in which tumor cells spread to new organs, is one of the greatest challenges in cancer research and is the leading cause of cancer-induced death. Numerous studies have shown that oncoviruses and their encoded proteins significantly affect metastasis, especially the EMT process. ALV-J and MDV are classic tumorigenic retrovirus and herpesvirus, respectively. We found that ALV-J and MDV synergistically promoted EMT. Further, we identified the tumor stem cell marker DCLK1 in ALV-J and MDV co-infected cells. DCLK1 directly interacted with ß-catenin, promoting the formation of the ß-catenin-TCF4 complex. This interaction activated the Wnt/ß-catenin pathway, thereby inducing EMT and paving the way for synergistic tumor metastasis. Exploring the molecular mechanisms by which ALV-J and MDV cooperate during EMT will contribute to our understanding of tumor progression and metastasis. This study provides new insights into the cooperative induced tumor metastasis by retroviruses and herpesviruses.

Demonstration of efficient Thomson cooler by electronic phase transition.

In the 1850s, Lord Kelvin predicted the existence of a thermoelectric cooling effect inside a whole material (the Thomson effect) according to thermodynamics1, in addition to the Peltier effect that enables cooling at the junction between dissimilar materials. However, the Thomson effect is usually negligible (ΔT/T < 2%) in conventional thermoelectric materials because the entropy change in charge carriers is fairly small2, leading to the guiding principles for advancing thermoelectric cooling to be based on the framework of the Peltier effect and that the figure of merit ZT should be maximized to optimize performance. Here, we demonstrate a Thomson-effect-enhanced thermoelectric cooler using a large Thomson coefficient (τ) induced by the direct manipulation of charge entropy through an electronic phase transition in YbInCu4. The devices achieve a steady temperature span (ΔT) of >5 K from T = 38 K. Our findings suggest not only another approach to advance thermoelectric coolers in addition to improving ZT but also technologically opens opportunities for solid-state cryogenic cooling applications.

The effector SJP3 interferes with pistil development by sustaining SHORT VEGETATIVE PHASE 3 expression in jujube.

Jujube witches' broom (JWB) is a phytoplasma disease that causes severe damage to jujube (Ziziphus jujuba) crops worldwide. Diseased jujube plants show enhanced vegetative growth after floral reversion, including leafy flower structures (phyllody) and the fourth whorl converting into a vegetative shoot. In previous research, secreted JWB protein 3 (SJP3) was identified as an inducer of phyllody. However, the molecular mechanisms of SJP3-mediated pistil reversion remain unknown. Here, the effector SJP3 was found to interact with the MADS-box protein SHORT VEGETATIVE PHASE 3 (ZjSVP3). ZjSVP3 was expressed in young leaves and during the initial flower bud differentiation of healthy jujube-bearing shoots but was constitutively expressed in JWB phytoplasma-infected flowers until the later stage of floral development. The SJP3 effector showed the same expression pattern in the diseased buds and promoted ZjSVP3 accumulation in SJP3 transgenic jujube calli. The N-terminal domains of ZjSVP3 contributed to its escape from protein degradation in the presence of SJP3. Heterologous expression of ZjSVP3 in Nicotiana benthamiana produced typical pistil abnormalities, including trichome-enriched style and stem-like structures within the leaf-like ovary, which were consistent with those in the mildly malformed lines overexpressing SJP3. Furthermore, ectopic expression of ZjSVP3 directly bound to the zinc finger protein 8 (ZjZFP8) and MADS-box gene SHATTERPROOF 1 (ZjSHP1) promoters to regulate their expression, resulting in abnormal pistil development. Overall, effector SJP3-mediated derepression of ZjSVP3 sustained its expression to interfere with pistil development, providing insight into the mechanisms of pistil reversion caused by JWB phytoplasma in specific perennial woody plant species.

Revealing the signaling of complement receptors C3aR and C5aR1 by anaphylatoxins.

The complement receptors C3aR and C5aR1, whose signaling is selectively activated by anaphylatoxins C3a and C5a, are important regulators of both innate and adaptive immune responses. Dysregulations of C3aR and C5aR1 signaling lead to multiple inflammatory disorders, including sepsis, asthma and acute respiratory distress syndrome. The mechanism underlying endogenous anaphylatoxin recognition and activation of C3aR and C5aR1 remains elusive. Here we reported the structures of C3a-bound C3aR and C5a-bound C5aR1 as well as an apo-C3aR structure. These structures, combined with mutagenesis analysis, reveal a conserved recognition pattern of anaphylatoxins to the complement receptors that is different from chemokine receptors, unique pocket topologies of C3aR and C5aR1 that mediate ligand selectivity, and a common mechanism of receptor activation. These results provide crucial insights into the molecular understanding of C3aR and C5aR1 signaling and structural templates for rational drug design for treating inflammation disorders.

Causal association among glaucoma, cerebral cortical structures, and Alzheimer's disease: insights from genetic correlation and Mendelian randomization.

Glaucoma and Alzheimer's disease are critical degenerative neuropathies with global impact. Previous studies have indicated that glaucomatous damage could extend beyond ocular structures, leading to brain alterations potentially associated with Alzheimer's disease risk. This study aimed to explore the causal associations among glaucoma, brain alterations, and Alzheimer's disease. We conducted a comprehensive investigation into the genetic correlation and causality between glaucoma, glaucoma endophenotypes, cerebral cortical surficial area and thickness, and Alzheimer's disease (including late-onset Alzheimer's disease, cognitive performance, and reaction time) using linkage disequilibrium score regression and Mendelian randomization. This study showed suggestive genetic correlations between glaucoma, cortical structures, and Alzheimer's disease. The genetically predicted all-caused glaucoma was nominally associated with a decreased risk of Alzheimer's disease (OR = 0.96, 95% CI: 0.93-0.99, P = 0.013). We found evidence for suggestive causality between glaucoma (endophenotypes) and 20 cortical regions and between 29 cortical regions and Alzheimer's disease (endophenotypes). Four cortical regions were causally associated with cognitive performance or reaction time at a significant threshold (P < 6.2E-04). Thirteen shared cortical regions between glaucoma (endophenotypes) and Alzheimer's disease (endophenotypes) were identified. Our findings complex causal relationships among glaucoma, cerebral cortical structures, and Alzheimer's disease. More studies are required to clarify the mediation effect of cortical alterations in the relationship between glaucoma and Alzheimer's disease.

Highly Potent Transparent Passive Cooling Coating via Microphase-Separated Hydrogel Combining Radiative and Evaporative Cooling.

Transparent passive cooling materials can cool targets environmentally without interfering with light transmission or visual information reception. They play a prominent role in solar cells and flexible display cooling. However, achieving potent transparent cooling remains challenging, because light transmission is accompanied by thermal energy. Here we propose to realize effective passive cooling in transparent materials via a microscale phase separation hydrogel film. The poly(N-isopropylacrylamide-co-acrylamide) hydrogel presents light transmittance of >96% and infrared emissivity as high as 95%. The microphase-separated structure affords a higher enthalpy of evaporation. The film is highly adhesive. In field applications, it reduces temperatures by 9.14 °C compared to those with uncovered photovoltaic panels and 7.68 °C compared to those for bare flexible light-emitting diode screens. Simulations indicate that energy savings of 32.76-51.65 MJ m-2 year-1 can be achieved in typical tropical monsoon climates and temperate continental climates. We expect this work to contribute to energy-efficient materials and a carbon-neutral society.

Identification and functional analysis of circulating small extracellular vesicle lncRNA signatures in children with fulminant myocarditis.

Fulminant myocarditis (FM) is the most serious type of myocarditis. However, the molecular mechanism underlying the pathogenesis of FM has not been fully elucidated. Small extracellular vesicles (sEVs) play important roles in many diseases, but any potential role in paediatric FM has not been reported. Here, the differential signatures of lncRNAs in plasma sEVs were studied in FM children and healthy children using transcriptome sequencing followed by functional analysis. Then immune-related lncRNAs were screened to study their role in immune mechanisms, the levels and clinical relevance of core immune-related lncRNAs were verified by qRT-PCR in a large sample size. Sixty-eight lncRNAs had increased levels of plasma sEVs in children with FM and 11 had decreased levels. Functional analysis showed that the sEVs-lncRNAs with different levels were mainly related to immunity, apoptosis and protein efflux. Seventeen core immune-related sEVs-lncRNAs were screened, functional enrichment analysis showed that these lncRNAs were closely related to immune activation, immune cell migration and cytokine pathway signal transduction. The results of the study show that sEVs-lncRNAs may play an important role in the pathogenesis of fulminant myocarditis in children, especially in the mechanism of immune regulation.

The distinct initiation sites and processing activities of TTLL4 and TTLL7 in glutamylation of brain tubulin.

Mammalian brain tubulins undergo a reversible posttranslational modification-polyglutamylation-which attaches a secondary polyglutamate chain to the primary sequence of proteins. Loss of its erasers can disrupt polyglutamylation homeostasis and cause neurodegeneration. Tubulin tyrosine ligase like 4 (TTLL4) and TTLL7 were known to modify tubulins, both with preference for the ß-isoform, but differently contribute to neurodegeneration. However, differences in their biochemical properties and functions remain largely unknown. Here, using an antibody-based method, we characterized the properties of a purified recombinant TTLL4 and confirmed its sole role as an initiator, unlike TTLL7, which both initiates and elongates the side chains. Unexpectedly, TTLL4 produced stronger glutamylation immunosignals for α-isoform than ß-isoform in brain tubulins. Contrarily, the recombinant TTLL7 raised comparable glutamylation immunoreactivity for two isoforms. Given the site selectivity of the glutamylation antibody, we analyzed modification sites of two enzymes. Tandem mass spectrometry analysis revealed their incompatible site selectivity on synthetic peptides mimicking carboxyl termini of α1- and ß2-tubulins and a recombinant tubulin. Particularly, in the recombinant α1A-tubulin, a novel region was found glutamylated by TTLL4 and TTLL7, that again at distinct sites. These results pinpoint different site specificities between two enzymes. Moreover, TTLL7 exhibits less efficiency to elongate microtubules premodified by TTLL4, suggesting possible regulation of TTLL7 elongation activity by TTLL4-initiated sites. Finally, we showed that kinesin behaves differentially on microtubules modified by two enzymes. This study underpins the different reactivity, site selectivity, and function of TTLL4 and TTLL7 on brain tubulins and sheds light on their distinct role in vivo.

The dorsomedial prefrontal cortex promotes self-control by inhibiting the egocentric perspective.

The dorsomedial prefrontal cortex (dmPFC) plays a crucial role in social cognitive functions, including perspective-taking. Although perspective-taking has been linked to self-control, the mechanism by which the dmPFC might facilitate self-control remains unclear. Using the multimodal neuroimaging dataset from the Human Connectome Project (Study 1, N =978 adults), we established a reliable association between the dmPFC and self-control, as measured by discounting rate-the tendency to prefer smaller, immediate rewards over larger, delayed ones. Experiments (Study 2, N = 36 adults) involving high-definition transcranial direct current stimulation showed that anodal stimulation of the dmPFC reduces the discounting of delayed rewards and decreases the congruency effect in egocentric but not allocentric perspective in the visual perspective-taking tasks. These findings suggest that the dmPFC promotes self-control by inhibiting the egocentric perspective, offering new insights into the neural underpinnings of self-control and perspective-taking, and opening new avenues for interventions targeting disorders characterized by impaired self-regulation.

Single-Site Pd Regulated by π-π Stacking for High-Selectivity Cyclopropanation Reaction.

Cyclopropane-based high-energy fuels possess high intramolecular energy and density, and their precise synthesis is a critical challenge. However, owing to the highest strain in the cyclopropane structure (compared to other four- or five-membered rings, etc.), metal-carbene intermediates form with difficulty, resulting in poor catalytic selectivity for its synthesis. Herein, through rational design of π-π stacking between the Pd organic complex and graphene, we report a single-site Pd catalyst for precise synthesis of multicyclopropane-based high-energy fuels. It is discovered that π-π stacking enhanced the electrophilicity of Pd through a weak metal-support interaction, thus promoting the formation of Pd═C carbene active intermediates. Meanwhile, the adsorption between the active centers and intermediates was enhanced via π-π stacking. These two respects led to almost twice selectivity for cyclopropanation reaction up to 80.5% as that without π-π stacking. This work provides an effective strategy of π-π noncovalent interactions for regulating C-C coupling reaction selectivity.

Unraveling the Transformation from Type-II to Z-Scheme in Perovskite-Based Heterostructures for Enhanced Photocatalytic CO2 Reduction.

The ability to create perovskite-based heterostructures with desirable charge transfer characteristics represents an important endeavor to render a set of perovskite materials and devices with tunable optoelectronic properties. However, due to similar material selection and band alignment in type-II and Z-scheme heterostructures, it remains challenging to obtain perovskite-based heterostructures with a favorable electron transfer pathway for photocatalysis. Herein, we report a robust tailoring of effective charge transfer pathway in perovskite-based heterostructures via a type-II to Z-scheme transformation for highly efficient and selective photocatalytic CO2 reduction. Specifically, CsPbBr3/TiO2 and CsPbBr3/Au/TiO2 heterostructures are synthesized and then investigated by ultrafast spectroscopy. Moreover, taking CsPbBr3/TiO2 and CsPbBr3/Au/TiO2 as examples, operando experiments and theoretical calculations confirm that the type-II heterostructure could be readily transformed into a Z-scheme heterostructure through establishing a low-resistance Ohmic contact, which indicates that a fast electron transfer pathway is crucial in Z-scheme construction, as further demonstrated by CsPbBr3/Ag/TiO2 and CsPbBr3/MoS2 heterostructures. In contrast to pristine CsPbBr3 and CsPbBr3/TiO2, the CsPbBr3/Au/TiO2 heterostructure exhibits 5.4- and 3.0-fold enhancement of electron consumption rate in photocatalytic CO2 reduction. DFT calculations and in situ diffuse reflectance infrared Fourier transform spectroscopy unveil that the superior CO selectivity is attributed to the lower energy of *CO desorption than that of hydrogenation to *HCO. This meticulous design sheds light on the modification of perovskite-based multifunctional materials and enlightens conscious optimization of semiconductor-based heterostructures with desirable charge transfer for catalysis and optoelectronic applications.

Unlocking Copper-Free Interfacial Asymmetric C-C Coupling for Ethylene Photosynthesis from CO2 and H2O.

Solar-driven carbon dioxide (CO2) reduction into C2+ products such as ethylene represents an enticing route toward achieving carbon neutrality. However, due to sluggish electron transfer and intricate C-C coupling, it remains challenging to achieve highly efficient and selective ethylene production from CO2 and H2O beyond capitalizing on Cu-based catalysts. Herein, we report a judicious design to attain asymmetric C-C coupling through interfacial defect-rendered tandem catalytic centers within a sulfur-vacancy-rich MoSx/Fe2O3 photocatalyst sheet, enabling a robust CO2 photoreduction to ethylene without the need for copper, noble metals, and sacrificial agents. Specifically, interfacial S vacancies induce adjacent under-coordinated S atoms to form Fe-S bonds as a rapid electron-transfer pathway for yielding a Z-scheme band alignment. Moreover, these S vacancies further modulate the strong coupling interaction to generate a nitrogenase-analogous Mo-Fe heteronuclear unit and induce the upward shift of the d-band center. This bioinspired interface structure effectively suppresses electrostatic repulsion between neighboring *CO and *COH intermediates via d-p hybridization, ultimately facilitating an asymmetric C-C coupling to achieve a remarkable solar-to-chemical efficiency of 0.565% with a superior selectivity of 84.9% for ethylene production. Further strengthened by MoSx/WO3, our design unveils a promising platform for optimizing interfacial electron transfer and offers a new option for C2+ synthesis from CO2 and H2O using copper-free and noble metal-free catalysts.

NR0B1 augments sorafenib resistance in hepatocellular carcinoma through promoting autophagy and inhibiting apoptosis.

NR0B1 is frequently activated in hepatocellular carcinoma (HCC). However, the role of NR0B1 is controversial in HCC. In this study, we observed that NR0B1 was an independent poor prognostic factor, negatively correlated with the overall survival of HCC and the relapse-free survival of patients treated with sorafenib. Meanwhile, NR0B1 promoted the proliferation, migration, and invasion of HCC cells, inhibited sorafenib-induced apoptosis, and elevated the IC50 of sorafenib in HCC cells. NR0B1 was further displayed to increase sorafenib-induced autophagic vesicles and activate Beclin1/LC3-II-dependent autophagy pathway. Finally, NR0B1 was revealed to transcriptionally suppress GSK3ß that restrains AMPK/mTOR-driven autophagy and increases BAX-mediated apoptosis. Collectively, our study uncovered that the ectopic expression of NR0B1 augmented sorafenib-resistance in HCC cells by activating autophagy and inhibiting apoptosis. Our findings supported that NR0B1 was a detrimental factor for HCC prognosis.

An Orthogonal CRISPR/dCas12a System for RNA Imaging in Live Cells.

CRISPR/Cas technology has made great progress in the field of live-cell imaging beyond genome editing. However, effective and easy-to-use CRISPR systems for labeling multiple RNAs of interest are still needed. Here, we engineered a CRISPR/dCas12a system that enables the specific recognition of the target RNA under the guidance of a PAM-presenting oligonucleotide (PAMmer) to mimic the PAM recognition mechanism for DNA substrates. We demonstrated the feasibility and specificity of this system for specifically visualizing endogenous mRNA. By leveraging dCas12a-mediated precursor CRISPR RNA (pre-crRNA) processing and the orthogonality of dCas12a from different bacteria, we further demonstrated the proposed system as a simple and versatile molecular toolkit for multiplexed imaging of different types of RNA transcripts in live cells with high specificity. This programmable dCas12a system not only broadens the RNA imaging toolbox but also facilitates diverse applications for RNA manipulation.

Enzymatically Mediated In Situ Generation of Z-Scheme Bi2S3/Bi2MoO6 Heterojunction-Based Organic Photoelectrochemical Transistor for METTL3/METTL14 Detection.

The OPECT biosensing platform, which connects optoelectronics and biological systems, offers significant amplification and more possibilities for research in biological applications. In this work, a homogeneous organic photoelectrochemical transistor (OPECT) biosensor based on a Bi2S3/Bi2MoO6 heterojunction was constructed to detect METTL3/METTL14 protein activity. The METTL3/METTL14 complex enzyme was used to catalyze adenine (A) on an RNA strand to m6A, protecting m6A-RNA from being cleaved by an E. coli toxin (MazF). Alkaline phosphatase (ALP) catalyzed the conversion of Na3SPO3 to H2S through an enzymatic reaction. Due to the adoption of the strategy of no fixation on the electrode, the generated H2S was easy to diffuse to the surface of the ITO electrode. The Bi2S3/Bi2MoO6 heterojunction was formed in situ through a chemical replacement reaction with Bi2MoO6, improving photoelectric conversion efficiency and realizing signal amplification. Based on this "signal on" mode, METTL3/METTL14 exhibited a wide linear range (0.00001-25 ng/µL) between protein concentration and photocurrent intensity with a limit of detection (LOD) of 7.8 fg/µL under optimal experimental conditions. The applicability of the developed method was evaluated by investigating the effect of four plasticizers on the activity of the METTL3/METTL14 protein, and the molecular modeling technique was employed to investigate the interaction between plasticizers and the protein.