Biology subjects

Wei, Q.

Publications and source records attributed to Wei, Q..

At least 19 recordsLinked to original sources

bioRxiv2026

Shared and distinct immunometabolic states link to divergent pathology in cutaneous mycobacterial infections

Cutaneous mycobacterial infections provide a common tissue setting in which related intracellular pathogens elicit distinct pathological responses, but the innate immune basis of this divergence remains incompletely understood. Here, we combined single-cell profiling from 33 individuals with histological and functional analyses, examining healthy skin and lesions from multibacillary leprosy (MB), paucibacillary leprosy (PB), cutaneous tuberculosis (CTB) and Mycobacterium marinum infection (Mari). All infections showed lower mononuclear phagocyte and higher natural killer (NK) cell proportions within the CD45 compartment, alongside shared and infection-associated dendritic cell, macrophage and NK cell programs. CTB and Mari displayed broader macrophage antimicrobial signatures, whereas MB showed lipid-associated programs and attenuated interferon-associated effector expression across multiple macrophage subsets. Reduced expression of fructose-1,6-bisphosphatase 1 (FBP1) was associated with the absence of Langhans giant cells in MB, and pharmacological FBP1 inhibition nearly abolished fusion of macrophages. Compared with NK cells from Mari, MB NK cells showed reduced central carbon and oxidative metabolism signatures alongside increased lipid-associated programs. Functional experiments showed that inhibition of glucose utilization, disruption of mitochondrial function, or co-culture with foamy macrophages abundant in MB lesions reduced NK cell IFN-{gamma} production. These findings identify shared and distinct innate immune programs and suggest that their interplay with local metabolic environments shapes divergent pathology across cutaneous mycobacterial infections.

immunology↗

bioRxiv2026

BCIJelly: An integrated ecosystem for brain-computer interface research

Brain-computer interface (BCI) research relies on multistage computational pipelines, but progress has been slowed by fragmented data formats, heterogeneous decoder implementations and hardware-specific deployment toolchains. Here, we introduce BCIJelly, a unified ecosystem that standardizes 18 BCI datasets into AI-ready inputs and integrates 15 benchmark decoders, 80 reusable modules, automated architecture search (AAS) and hardware-aware neuromorphic deployment. Our AAS constructs task-specific decoders without manual design and extends into a large language model (LLM)-driven closed-loop mode supporting single-task, multitask and cross-species decoder design. A single-command pipeline compiles trained decoders for neuromorphic hardware, reducing power consumption by 30 to 50 times while preserving decoding performance. An interactive visualization software enables code-free exploration of neural recordings and decoding outputs. BCIJelly is validated across five BCI paradigms (motor, visual, speech, emotion and auditory) in humans, macaques and mice, providing an extensible ecosystem connecting data standardization, decoder development, systematic evaluation and hardware-aware deployment for BCI research.

neuroscience↗

bioRxiv2026

A Quantitative Two-Channel Genetic Reporter for Selenocysteine Biosynthesis and Incorporation

Selenocysteine (Sec), the 21st amino acid, is a rare non-canonical amino acid that represents an attractive target for protein engineering due to its desirable chemical properties such as high affinity for metals, strong nucleophilicity, and reversible covalent bond formation. To bypass the natural constraints on Sec placement within proteins, several strategies have been developed to rewire the native translational machinery to enable site-specific incorporation. However, these usually abolish the quality control mechanism that excludes the serine-charged selenocysteinyl-tRNA (Ser-tRNASec), the immediate biosynthetic precursor, from translation resulting in heterogenous protein species. This challenge is confounded by a lack of genetic tools to accurately report the selenylation state of the tRNA pool as most are blind to competing process of Ser incorporation, which can only be observed using analytical methods. To resolve this issue, we have developed a new fluorescent reporter, Selenocysteine Adjusted Ratiometric Chromophore (SeARCh), which exhibits two distinct spectral outputs dependent on the incorporation of either Ser (red) or Sec (green). Using SeARCh, we define several factors which influence the observed Sec:Ser ratio and construct a new hybrid biosynthetic pathway with improved performance, achieving 90% Sec incorporation. Furthermore, SeARCh displays unusually complex mass spectra due to the isotope distribution of selenium and heterogenous nature of the protein in solution and we report specific methods to account for this behaviour and precisely quantify the rare Ser-containing species found at high Sec incorporation efficiencies. Our findings suggest that the equilibrium between selenoprotein and tRNASec expression levels is a key driver of incorporation efficiency and implies a process that is broadly biosynthetically constrained. Collectively these tools represent a significant advance in the metrology of selenocysteine biosynthesis and incorporation and can be used to inform and standardize future engineering efforts.

synthetic biology↗

bioRxiv2026

Discrete depolarisations of chordotonal organ neurons propagate toward the soma and are independent of the sensory TRP channels NompC and Nanchung-Inactive.

The electrophysiological process of auditory transduction in insects remains largely conjecture due to the unknown role of ion channels localised to the cilia, but experimental evidence supports either NompC or Nan-Iav as the auditory mechanotransduction ion channel. Here, we knocked down two key genes that code for the two candidate sound-activated ion channels using dsRNA-mediated RNA interference. We measured sound-evoked activity of the auditory nerve and intracellular electrical currents from the ciliated ending of individual auditory receptors. We found that the sound-evoked nerve activity was reduced in nompC, nan and ift88 knockdown. Using whole-cell patch clamp recordings we found that nompC and nan knockdown resulted in reduced sound-evoked transduction current. Stochastic depolarisations hypothesised to be mediated from one of the candidate mechanotransduction ion channels, either NompC or Nan-Iav, where not affected by knockdown of either channel. The discrete depolarisations are therefore mediated through another unidentified ion channel. We test the hypothesis that discrete depolarisations are graded action potentials that travel toward the soma through noise analysis of the transduction current and analysis of discrete depolarisations to voltage-steps. As a positive control we also knocked down ift88, a protein essential for transporting proteins, including ion channels, along the cilium and found both the transduction current and the discrete depolarisations decreased. Key pointsO_LIInjection of dsRNA decreased RNA of nompC and nan C_LIO_LISound-evoked nerve activity is reduced for RNAi-mediated knockdown of nompC and nan C_LIO_LINompC and Nan both contribute to the transduction current C_LIO_LIThe stochastic discrete depolarisations are not due to NompC or Nan-Iav ion channel but to a third unidentified ion channel. C_LIO_LINoise analysis of the transduction current and the discrete depolarisations suggests they are graded action potentials that travel in the direction of the soma. C_LIO_LIKnockdown of ift88 reduced both the transduction current and discrete depolarisations. C_LI Significance StatementInsects are important to understand, economically, agriculturally and medically. However, we still do not understand fundamental aspects of how insects detect their own body movements, vibrations and sound. These senses are detected by insect chordotonal organs, specialised miniaturised mechanoreceptors that convert movements into electrical signals through specialised ion channels. Previous experimental work has advocated either NompC or Nan-Iav as the mechanosensitive ion channel. Here, for the first time, we reduced the expression of both nompC and nan and measured the sound-evoked transduction current directly from neurons in a specialised auditory chordotonal organ. In contradiction to previous studies, we show that both ion channels contribute to the transduction current and find that electrical signals termed "discrete depolarisations" travel toward the soma.

neuroscience↗

bioRxiv2026

The Fas-FADD-caspase-8 axis is a cancer cell-intrinsic determinant of cytotoxic lymphocyte-mediated killing

Cytotoxic lymphocytes induce cancer cell death through death receptor-ligand interactions and the perforin-granzyme pathway. These pathways are generally thought to converge on the activation of executioner caspases to drive apoptosis. Here, we employed a reductionist approach to systematically disrupt key cell death mediators in a cytotoxic lymphocyte killing system to define their roles in determining cancer cell fate. We found that loss of executioner caspases conferred only limited resistance to cytotoxic lymphocyte-mediated killing. To identify cancer cell-intrinsic regulators that function beyond executioner caspases, we performed unbiased genome-wide CRISPR screens in executioner caspase-deficient cells. Unexpectedly, disruption of Fas or FADD--core components of the death receptor pathway-- conferred substantial resistance to cytotoxic lymphocyte-mediated killing even in the absence of executioner caspases. This resistance persisted following additional disruption of known downstream mediators of Fas-FADD-caspase-8 (CASP8) signaling. Together, these findings identify the Fas-FADD-CASP8 axis as a central cancer cell-intrinsic determinant of susceptibility to cytotoxic lymphocyte-mediated killing whose function is not fully explained by canonical apoptotic or non-apoptotic effector pathways. Our results further suggest that CASP8 engages additional downstream substrates or mechanisms to promote cytotoxic lymphocyte-induced cancer cell death.

immunology↗

bioRxiv2026

Rhythmic temporal structure organizes recurrent dynamics to support sequential working memory

Rhythmic temporal structure improves working memory, but how this benefit emerges from recurrent dynamics remains unclear. Here, we trained excitatory-inhibitory recurrent neural networks with short-term synaptic plasticity to perform a sequential delayed match-to-sample task with either regular or jittered sample timing. Rhythmic input produced a small but reliable improvement in task accuracy and was associated with more differentiated population trajectories during encoding. This behavioral advantage was accompanied by an organization of population dynamics around the dominant input frequency: temporal regularity progressively brought stimulus arrivals closer to preferred encoding phases, modulated phase advancement during stimulus presentation, and reduced the deviation of inter-stimulus phase-progression frequency from the dominant input rhythm. As a result, internal oscillations increasingly tracked the temporal structure of the input across the sequence, providing a phase-based scaffold for encoding ordered information. This scaffold preferentially supported temporal-order representations rather than uniformly enhancing all stimulus features. Decoding analyses further showed that stronger temporal regularity increased the fidelity and persistence of stimulus information in both neuronal activity and synaptic efficacy, whereas perturbing synaptic efficacy produced the largest impairment during delay-period maintenance. These findings suggest that rhythmic input supports sequential working memory by imposing a reliable temporal structure on recurrent dynamics and stabilizing synaptic-state representations.

neuroscience↗

bioRxiv2026

Computational and Proteomic Analyses Reveal Cardiac Dysfunction and Heart Failure-Associated Biomarker Secretion from Venezuelan Equine Encephalitis Virus TC83-infected human IPSC-derived Cardiomyocytes

Arthropod-borne pathogens, many of which are neurotropic, can disseminate beyond the central nervous system to infect peripheral organs. In recent years, an increasing number of cardiac dysfunctions have been reported following arthropod-borne viral infections; however, the mechanism underlying these cardiac manifestations remains poorly understood. In this study, we investigated the impact of Venezuelan Equine Encephalitis Virus (VEEV) TC-83 infection on cardiac function and immune-response of human induced-pluripotent stem cell (hIPSC)-derived cardiomyocytes (hIPSC-CMs). We first confirmed the successful differentiation of hIPSCs into spontaneously beating hIPSC-CMs. We then demonstrated that these cells are highly susceptible to VEEV TC-83 infection, which induced pronounced arrhythmias and complete cessation of beating within 24 hours post-infection. To quantify these functional changes, we developed a segmentation-free computational pipeline that converts frame-to-frame motion in brightfield time-lapse movies into a one-dimensional signal reflecting contractile activity and extracts beat timing, beat rate, and rhythm-regularity features in the time and frequency domains. This analysis revealed progressive disruption of beating dynamics following VEEV TC-83 infection, with early rhythm instability and complete loss of coordinated beating by 24 hours post-infection. Furthermore, mass spectrometry analysis of VEEV TC-83-infected hIPSC-CMs supernatants revealed the presence of biomarkers typically associated with heart failure in patients, underscoring a virus-induced cardiac functional impairment. Together, these findings provide new insight into cardiac complications associated with arthropod-borne viral infections and may support advances in preventive medicine.

cell biology↗

bioRxiv2026

Hyperactive STAT1 Promotes T Follicular Helper Type 1 Cell Differentiation to Trigger Autoimmunity

ABSTRACTSHeterozygous gain-of-function (GOF) mutations in signal transducer and activator of transcription 1 (STAT1) cause an inborn error of immunity characterized by immune dysregulation, recurrent infections and various autoimmune manifestations. However, the precise pathogenic mechanism by which STAT1 GOF contributes to autoimmunity remains elusive. In our cohort, STAT1-GOF patients exhibit biased circulating follicular helper T (cTfh) populations with CXCR3+ Tfh1-like features. Using a Stat1 GOF mouse model that spontaneously developed autoimmunity, we found that overactivated STAT1 promotes Tfh differentiation and disrupted T cell-dependent humoral responses with skewed immunoglobulin class switching towards IgG2. Furthermore, STAT1 GOF directly targets to Tfh and Th1 cell signature genes and thereby drives the development of Tfh1 cells with excessive IFN-{gamma} production, which implicated in autoantibody production and the development of autoimmunity. Notably, IFN-{gamma} neutralization significantly alleviated autoimmune cellular responses and autoantibody levels in mutant mice, highlighting IFN-{gamma} blockade as a promising targeted therapy for the STAT1-GOF patients with autoimmunity. Our findings suggest that proper regulation of STAT1 activity within a reasonable magnitude is crucial for ensuring optimal host-protective humoral immunity. One-sentence summaryOveractivated STAT1 promotes Tfh1 differentiation to drive autoimmunity.

immunology↗

bioRxiv2026

The neural control of infanticide and parental behaviors in male mice

Infanticide, the killing of conspecific young, is a natural behavior observed commonly in non-parental animals across species, including mice. Our recent study in female mice revealed a mutually inhibitory circuit, composed of estrogen receptor alpha cells in the medial preoptic area (MPOAEsr1) and the posterior part of the bed nucleus of stria terminalis (BNSTpEsr1), that controls pup-directed behaviors, with the former driving maternal care and the latter promoting infanticide. Given that both MPOA and BNSTp are sexually dimorphic, here we asked whether the same circuit operates in males. Our functional manipulations and in vivo and in vitro recordings reveal that MPOAEsr1 and BNSTpEsr1 cells similarly and respectively drive paternal care and infanticide and antagonize each other in male mice. Furthermore, during fatherhood, MPOAEsr1 cell excitability increases while BNSTpEsr1 cell excitability decreases to enable the switch from infanticide to paternal care. Despite the similarity in circuit organization, a direct comparison between males and females reveals sex differences in the intrinsic properties of MPOAEsr1 and BNSTpEsr1 cells. Thus, MPOAEsr1-BNSTpEsr1 emerges as a common circuit motif for controlling pup-directed behaviors in both sexes, whereas the activity balance between these two populations differs between sexes and likely contributes to the different tendencies in males and females to express pup-caring versus killing behaviors.

neuroscience↗

bioRxiv2025

Mapping the phenotypic landscape of a transcriptional repressor using Deep Mutational Scanning and Growth-based Quantitative Sequencing

CymR is a TetR-family transcriptional repressor that recognizes a well-defined operator sequence in the promoter PcymRC. The native ligand cumate and several structurally related aromatic acids bind at an allosteric site and induce a conformational change in CymR, resulting in release from the DNA operator and de-repression of the promoter. The amino acid residues that contribute to these core functions have not been mapped, nor has the protein been subjected to extensive mutagenesis to modify its function. Here, for the first time, we integrate Deep Mutational Scanning (DMS) with Growth-based Quantitative Sequencing (GROQ-Seq) to evaluate a comprehensive phenotypic landscape of CymR variants, including single amino acid insertions and deletions. We measure this library across a concentration gradient of small molecule inducers to construct an induction curve for all library members. From this analysis, we identify amino acids throughout the protein that are essential for repressor function and discover several mutations that improve the sensitivity of CymR to the ligand perillic acid. In addition, rarely investigated insertion mutants are revealed to be a key driver of novel phenotypes, including several regions of CymR where insertions result in an inverted phenotype and the isolation of variants exhibiting an unusual band-stop phenotype. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=81 SRC="FIGDIR/small/693801v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@121b16org.highwire.dtl.DTLVardef@b0478aorg.highwire.dtl.DTLVardef@128f2ecorg.highwire.dtl.DTLVardef@164a426_HPS_FORMAT_FIGEXP M_FIG C_FIG

synthetic biology↗

bioRxiv2025

Engineering a Bright Near-Infrared Fluorescent Protein by Screening a Comprehensive Phenotypic Landscape

Near-infrared fluorescent proteins (niRFPs) are valuable markers for tracking cellular phenomena as they offer greater imaging depth, lower background, and minimal invasiveness relative to other fluorescent probes. The small ultra red fluorescent protein (smURFP) is the brightest niRFP currently reported and has been the subject of several mutagenesis studies to improve its biophysical characteristics. Here, we demonstrate a systematic approach to exploring the mutational landscape of smURFP using a comprehensive deep mutational scanning (DMS) library to identify novel smURFP mutations which confer greater in vivo fluorescence. By observing changes in relative abundance between naive and fluorescence sorted populations, we provide analysis of the enrichment of all possible single-codon substitutions, insertions, and deletions for smURFP. Enriched populations yielded a series of seven single-codon substitutions which confer a three-fold increase in in vivo fluorescence in E. coli when combined. Finally, we assess the potential underlying mechanisms for increased fluorescence by characterizing the biophysical and photophysical properties of the mutant niRFP sequences. We confirm that two of the derived smURFP sequences have a higher molecular brightness than wild type and yield the brightest niRFP reported.

synthetic biology↗

bioRxiv2025

Multi-strain Analysis of Pseudomonas putida Reveals the Metabolic and Genetic Diversity of the Species.

Pseudomonas putida is a gram-negative bacterial species increasingly utilized in biotechnology due to its robust growth, ability to degrade aromatic compounds, solvent tolerance, and genetic tractability. In this study, we report a comprehensive multi-strain analysis of 164 P. putida strains. We performed whole-genome sequencing and hybrid assembly for 40 strains, contributing a [~]8% increase to the available genomic data for P. putida. Furthermore, high-throughput phenotypic profiling using the Biolog phenotype microarray system for 24 strains on 190 unique carbon sources, along with 15 aromatic compounds not present on Biolog plates, yielded 4,920 unique strain-phenotype measurements. These data were leveraged to curate GEMs for 24 representative strains, including a refined model for strain KT2440, which comprised 1,480 genes and 2,191 metabolites, achieving a prediction accuracy of 91.2% in carbon utilization. Systematic comparison of genomes and GEMs revealed both conserved core pathways and significant allelic and functional divergence across strains, highlighting strain-specific variation in aromatic degradation. While pathways for protocatechuate and phenylacetate degradation were widely conserved, metabolic capabilities for compounds such as ferulate, phenol, and cresols varied markedly, suggesting adaptation to distinct ecological niches. Alleleome analysis of enzymes such as PcaI and PcaJ revealed distinct, functionally similar clades, indicating possible convergent evolution or horizontal gene transfer. These results provide computable resources and models for selecting P. putida strains with desired traits for biomanufacturing and bioremediation and offer insights into the evolution and phylogeny of the P. putida species.

bioengineering↗

bioRxiv2025

Identification of a broad and potent V3 glycan site bNAb targeting anN332gp120 glycan-independent epitope

Broadly neutralizing antibodies (bNAbs) against HIV-1 can suppress viremia in vivo and inform vaccine development. Here, we characterized 007, a V3 glycan site bNAb exhibiting high levels of antiviral activity against multiclade pseudovirus panels1-3 (GeoMean IC50 = 0.012 {micro}g/mL, breadth = 69%, 217 virus strains) by targeting a N332gp120 glycan-independent V3 epitope, a site of Env vulnerability to which only weakly neutralizing antibodies had previously been identified. Functional analyses demonstrated distinct binding and neutralization profiles compared to classical V3 glycan site bNAbs. A 007 Fab-Env cryo-EM structure revealed contacts with the V3 324GD/NIR327 motif and interactions with N156gp120 and N301gp120 glycans. In contrast to classical V3 bNAbs, 007 binding to Env does not depend on the N332gp120 glycan, rendering it resistant to common escape mutations. Structures of 007 IgG-Env trimer complexes showed two Env trimers crosslinked by three bivalent IgGs, and bivalent 007 IgG was up to [~]300-fold more potent than monovalent 007 IgG heterodimer, suggesting a role for avidity in potent neutralization. Finally, in HIV-1ADA-infected humanized mice, 007 caused transient decline of viremia and overcame classical V3 escape mutations, highlighting 007s potential for HIV-1 prevention, therapy, functional cure, and vaccine design.

immunology↗

bioRxiv2025

Post-Translational Modifications Remodel Proteome-Wide Ligandability

Post-translational modifications (PTMs) vastly expand the diversity of human proteome, dynamically reshaping protein activity, interactions, and localization in response to environmental, pharmacologic, and disease-associated cues. While it is well established that PTMs modulate protein function, structure, and biomolecular interactions, their proteome-wide impact on small-molecule recognition--and thus druggability--remains largely unexplored. Here, we introduce a chemical proteomic strategy to delineate how PTM states remodel protein ligandability in human cells. By deploying broad profiling photoaffinity probes, we identified over 400 functionally diverse proteins whose ability to engage small molecules is impacted by phosphorylation or N-linked glycosylation status. Integration of binding site mapping with structural analyses revealed a diverse array of PTM-dependent pockets. Among these targets, we discovered that the phosphorylation status of common oncogenic KRAS mutants impact the action of small molecules, including clinically approved inhibitors. These findings illuminate an underappreciated, PTM-governed layer of proteome plasticity and uncover opportunities for the development of chemical probes to selectively target proteins in defined modification states.

biochemistry↗

bioRxiv2025

Dual Role of Glycosylation in Resistance to CD4-binding Site Broadly Neutralizing Antibodies

Broadly neutralizing antibodies (bNAbs) provide a useful tool for HIV cure strategies because of their ability to target conserved regions on the envelope (Env) protein in the context of both virions and infected cells. One of the most well studied bNAbs is the CD4 binding site (CD4bs) antibody, VRC01 and others in its class. A major obstacle to effective cure strategies with bNAbs is viral immune escape. A deeper understanding of escape pathways from VRC01-class antibodies in genetically diverse samples is needed. Using an in vitro viral escape assay where infected CD4+ T cells were cultured in the presence of increasing VRC01 concentrations, complete resistance to VRC01 was detected in Env 246.F3 by day 42. We determined that resistance was due to a mutation at position N276 that resulted in elimination of the glycan-attachment site. As the loss of a glycan at this site is known to increase virus sensitivity to CD4bs antibodies, we explored this finding further. Specifically, we introduced N276D mutation into the 12-virus global Env panel and measured neutralization susceptibility to a panel of CD4bs bNAbs. This N276D mutation increased resistance or sensitivity depending on the Env and bNAb in question, emphasizing a dual role of glycan N276 in VRC01-class bNAb neutralization. The role of this glycan in escape was demonstrated to be dependent on the context of the Env, the bNAb and the glycosylation complexity of the virus due to producer cells. These findings underscore the complexity of glycosylation in genetically diverse HIV escape from antibodies. ImportanceDespite decades of research, an HIV cure remains elusive, largely due to the viruss immense genetic variability and ability to evade immune clearance. While antiretroviral therapy (ART) suppresses viral replication, it does not eradicate the virus and presents long-term challenges related to toxicity, access, and adherence. Broadly neutralizing antibodies (bNAbs), particularly those targeting the conserved CD4 binding site such as VRC01 and others in its class, offer promise for durable control or cure by targeting both circulating virus and infected cells. However, viral escape from bNAbs remains a critical hurdle. In this study, we demonstrate that glycan-mediated escape from VRC01-class bNAbs is highly context-dependent--shaped by Env, bNAb, and the glycosylation patterns introduced by the producer cell. These findings emphasize the dual role of glycans in affecting antibody sensitivity and underscore the importance of viral and host factors in shaping effective bNAb-based cure strategies across diverse HIV-1 strains.

immunology↗

bioRxiv2025

Directed evolution of L-DOPA sensing and production enables efficient incorporation of an expanded genetic code into proteins

3,4-dihydroxyphenylalanine (L-DOPA) is a catechol-containing amino acid derived from L-tyrosine that can be introduced into proteins primarily as a post-translational modification. It can also be introduced in a site-specific manner into proteins via orthogonal tRNA synthetase:tRNA pairs. However, despite the interest in expanded genetic codes containing L-DOPA, efforts to improve metabolism relating to its production have been limited. Here, we report and engineer a LysR-family transcription factor, PP_2251, that can serve as a biosensor for L-DOPA, thereby allowing the optimization of genetic circuits for vastly improved L-DOPA production via compartmentalized partnered replication (CPR), an emulsion PCR-based directed evolution scheme. Using this biosensor, a promiscuous E. coli flavin-dependent monooxygenase (HpaB) was optimized via CPR for enhanced L-DOPA biosynthesis. Engineered HpaB variants could support efficient translational incorporation of L-DOPA into proteins with 80% incorporation efficiency and yields in excess of 250 mg.L-1. Improvements in L-DOPA production are not only of great interest for protein engineering and genetic code expansion but should impact the production of DOPA-derived natural products including catecholamine neurotransmitters and benzylisoquinoline alkaloids (BIAs), molecules of intense pharmaceutical interest.

synthetic biology↗

bioRxiv2025

Attomolar Detection of HIV-1 with Label-Free RCA-rCRISPR on Smartphone

Human Immunodeficiency Virus-1 (HIV) remains a major global public health challenge, having led to over 42.3 million deaths since its discovery in the early 1980s. Despite progress in prevention and treatment, around 60% of people with HIV (PWH) remain undiagnosed in resource-limited regions, disproportionately affecting vulnerable populations and underserved communities across the world. This illustrates the critical need for accessible, accurate, and equipment-free diagnostic tools to enhance detection and thus provide opportunities to curb its spread. Here, we developed a low-cost, robust, and label-free rolling circle amplification (RCA)-rCRISPR diagnostic platform for detecting HIV viral load with minimal instrumentation. Our strategy, combining the integration of RNA-detecting RCA reaction with plasmid reporter-based ratiometric CRISPR (rCRISPR), enables sensitive detection of unprocessed RNA targets without the need for intensive sample pre-treatment. This label-free RCA-rCRISPR diagnostic platform detected HIV RNA down to single-digit aM sensitivity (~3000 copies/mL) from PWH-derived HIV samples ex vivo. Unlike typical RCA, which requires sample fragmentations to break long RNA target sequences, our design harnesses the triple functions of the phi29 DNA polymerase (namely exonuclease activity, polymerization, and strand displacement), enabling the detection of the entire HIV genome without pre-fragmentation. For point-of-care (POC) applications, we constructed an all-in-one smartphone-based minigel electrophoresis device to facilitate equipment-free HIV viral load testing, making it accessible to resource-limited communities. Additionally, the assay has demonstrated the ability for point mutation detection (BRAF mutation in canine urothelial carcinoma), showcasing the robustness of our strategy for broad disease diagnostic applications.

bioengineering↗

bioRxiv2025

Microneedle-based precision payload delivery in plants

Traditional crop delivery methods, such as foliar spray and soil application, face significant limitations, including nutrient loss, environmental impacts, and low delivery efficiency. Recent advances in nanomaterials have offered novel molecular delivery platforms, but challenges such as synthesis complexity, long-term stability, and compliance with rigorous biosafety regulations persist. To provide a simpler, lower-cost, and safer alternative, we developed a polyvinyl alcohol (PVA)-based microneedle (MN) delivery system that can be precisely applied to various plant tissues (e.g., stem, lateral branch, or petiole), which demonstrates high delivery efficiency compared to the conventional methods (3.5x higher tissue accumulation) while reducing application dose (>90% less). This MN system facilitates the delivery of diverse small molecules, ranging from fluorescent dyes, growth promoters, to antiviral hormones, into plant tissues, on the other hand showing limited wounding stress to the plant. By applying fluorescent dye-loaded MNs onto tomato stems, we demonstrated effective molecular diffusion through vascular tissues. Additionally, MNs loaded with gibberellic acid (GA3) enhanced stem and branch growth in tomatoes and restored the lateral flowering phenotype in Arabidopsis ft-10 mutants, with significant upregulation of GA receptor gene expression. Lastly, salicylic acid (SA) injections with MNs induced resistance to tomato spotted wilt virus (TSWV) in Nicotiana benthamiana, comparable to conventional spray and infiltration-based approaches. This easily fabricated and cost-effective MN system offers a promising tool for precision agriculture, enhancing plant health and productivity while significantly reducing the use of agrochemicals.

plant biology↗