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Kumar, S.

Publications and source records attributed to Kumar, S..

At least 91 records · Page 5Linked to original sources

First haplotype-resolved genome assembly of citral-rich lemongrass Cymbopogon flexuosus var. Krishna

Cymbopogon flexuosus var. Krishna (lemongrass) is an aromatic grass valued for its high citral content, which is widely used in the fragrance, flavor, and pharmaceutical industries. C. flexuosus, a member of the Poaceae family, is a predominantly outcrossing species characterized by a highly heterozygous genome. Despite its economic importance and widespread cultivation, a high-quality reference genome has been lacking. Here, we report the first chromosome-scale genome assembly of lemongrass, generated using PacBio HiFi long-read sequencing combined with Omni-C chromatin conformation capture data. The resulting pseudo-haploid assembly spans approximately 798 Mb, organized into 10 chromosomes, and exhibits a scaffold N50 of 64.35 Mb. The assembly demonstrates high completeness, with 99.8% BUSCO recovery, and comprises [~]37,254 predicted protein-coding genes. In addition, we generated haplotype-resolved assemblies that capture the allelic diversity of this heterozygous genome. The haplotypes have sizes of [~]750 Mb and [~]726 Mb, representing 95-98% of the pseudo-haploid genome, and together they provide phase-resolved information for gene families and biosynthetic pathways. These high-quality assemblies establish a foundational genomic resource for advancing molecular breeding, comparative genomics, and metabolic engineering of lemongrass and related aromatic grasses.

genomics↗

Structural Characterization of the Type IV Secretion System in Brucella melitensis for Virtual Screening-Based Therapeutic Targeting

Brucellosis is a globally important zoonotic disease caused by Brucella melitensis, the most virulent and clinically significant species affecting both humans and livestock. Unlike many Gram-negative pathogens, B. melitensis, a facultative intracellular pathogen, lacks conventional virulence factors and instead relies on specialized systems such as the Type IV Secretion System (T4SS) for secretion of effector proteins. In this study, an integrated computational pipeline was implemented to identify, model, and assemble the T4SS components, encoded by virB operon, from the complete B. melitensis proteome. Template-based modeling strategies were employed to generate structures of T4SS subcomplexes, referencing crystallographic data from E. coli T4SS. Structural superposition with E. coli homologs revealed highly conserved architecture despite only 30-50% sequence identity. Stereochemical validation confirmed high model quality and favorable interactions among most VirB protein pairs. Membrane insertion analysis of the membrane-embedded assemblies further corroborated the spatial orientation of the modeled T4SS. Potential of T4SS as a drug target was explored by targeting dimeric interface of VirB11 ATPase to disrupt protein-protein interactions that could disarm the pathogen. Virtual screening of compounds from DrugBank database revealed compounds with docking score [≤] -7.0 kcal/mol that were screened based on ADMET properties, yielding three promising candidates - Ezetimibe (Drug Id: DB00973), Chlordiazepoxide (Drug Id: DB00475), and Alloin (Drug Id: DB15477). MM-GBSA analysis estimated favorable binding free energies for these compounds and molecular dynamics simulation for 200 ns further confirmed the protein-ligand interaction stability. Collectively, these findings provide new insights into the architecture of B. melitensis T4SS and identify three potential drug molecules targeting T4SS. This supports FDA - approved drug repurposing as an effective strategy for anti-virulence therapy against Brucellosis. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=103 SRC="FIGDIR/small/706537v1_ufig1.gif" ALT="Figure 1"> View larger version (46K): org.highwire.dtl.DTLVardef@847c4borg.highwire.dtl.DTLVardef@1fc2551org.highwire.dtl.DTLVardef@f62a7corg.highwire.dtl.DTLVardef@15f3468_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

FiCOPS: Hardware/Software Co-Design of FPGA Computational Framework for Mass Spectrometry-Based Peptide Database Search

Improving the speed and efficiency of database search algorithms that deduce peptides from mass spectrometry (MS) data has been an active area of research for more than three decades. The significance of the need for faster database search methods has rapidly increased due to the growing interest in studying non-model organisms, meta-proteomics, and proteogenomic data, which are notorious for their enormous search space. Poor scalability of serial algorithms with the growing size of the database and increasing parameters of post-translational modifications is a widely recognized problem. While high-performance computing techniques can be used on supercomputing machines, the need for real-time, on-the-instrument solutions necessitates the development of an efficient sytem-on-chip that optimizes design constraints such as cost, performance, and power of the system. To show case that such a system can work, we present an FPGA-based computational framework called FiCOPS to accelerate database search using a hardware/software co-design methodology. First, we theoretically analyze the database-search algorithm (closed-search) to reveal opportunities for parallelism and uncover computational bottlenecks. We then design an FPGA-based architectural template to exploit parallelism inherent in the search workload. We also formulate an analytical performance model for the architecture template to perform rapid design space exploration and find a near-optimal accelerator configuration. Finally, we implement our design on the Intel Stratix 10 FPGA platform and evaluate it using real-world datasets. Our experiments demonstrate that FiCOPS achieves 3.5 x speed-up over existing CPU solutions and 3x and 5x reduction in power consumption compared to existing CPU and GPU solutions.

bioinformatics↗

Neotaphonomic characteristics of vertebrate site formation in underwater caves

Recovering well-preserved vertebrate remains from underwater caves has provided critical insights into archaeological and palaeontological records worldwide. However, understanding how bone assemblages form and are modified in underwater environments remains limited due to stable low energy burial conditions that produce time-averaged deposits, and underwater settings that hinder traditional recording and recovery methods. This study applies an actualistic taphonomic framework to three assemblages of domesticate animal bones (N = 231) from two underwater caves, Green Waterhole and Gouldens Sinkhole, near Mount Gambier, South Australia, encompassing known submerged (wet; N = 134) and dry (N = 97) burial conditions. The assemblages were examined to assess how wet and dry cave environments impact bone distribution, surface and microstructural modification. Radiocarbon dating of 41 specimens indicates that domesticate fauna were deposited over decadal and centennial timescales, allowing taphonomic signatures to be contextualised through time. Statistically significant differences were identified between wet and dry burial contexts. Bones recovered from wet contexts exhibit mostly better preservation, including skeletal elemental completeness, surface, and microstructure, than those from dry caves. However, some of the submerged specimens also have elevated frequencies of bone surface corrosion with macroscopic evidence for heterogenous black biological staining, algal or biofilm attack, and a distinctive form of circular etching. Histotaphonomy further reveals patterns of peripheral cyanobacterial tunnelling across most bones recovered from submerged contexts. Bones from dry environments were dominated by terrestrially linked tunnelling across all regions of the bone cortex. These findings can be explained by variation in light availability across different cave zones which influences biological activity and, in turn, the expression of taphonomic markers on bone externally and at the microstructural level. This is the first study to provide a benchmark bone dataset for reconstructing depositional histories and post-depositional reworking in underwater cave environments under a taphonomic framework.

paleontology↗

Annelid eye evolution revealed by developmental, ultrastructural, and connectome analyses of cerebral eyes in Malacoceros fuliginosus

Eye evolution has long attracted interest, yet how multiple cerebral eyes within a lineage originate and diversify remains unclear. Annelids display exceptional diversity in eye number and structure, but the homology and function of distinct eye pairs are poorly understood. Here we investigate the cerebral eyes of the sedentary annelid Malacoceros fuliginosus using an integrated developmental, molecular, ultrastructural, and connectomic approach. We show that both the early-developing ventral and later-developing dorsal eyes are simple, few-celled, inverted rhabdomeric eyes that express transcription factors with conserved roles in animal eye development. Two r-opsin paralogs and distinct neurotransmitters are differentially expressed in different photoreceptor cells of the eyes. Ultrastructural reconstructions across larval development reveal differences in cellular composition and growth dynamics, while axonal tracing shows that photoreceptors from ventral and dorsal eyes project to overlapping regions of the larval brain. The overall organization and projections resemble those described in the errant annelid Platynereis dumerilii. Together, these data support the hypothesis that an ancestral cerebral eye duplicated early in annelid evolution, giving rise to multiple eye pairs with stage-specific functions.

evolutionary biology↗

A First-In-Class Broad Spectrum Inhibitor of Copper Exporting P1B-type ATPases

Copper (Cu) transporting ATPases represent a highly conserved subclass of P-type ATPases with critical roles in Cu export and metalloenzyme synthesis. Despite their important biological roles and association with a wide range of human diseases, no high-affinity small-molecule inhibitors have been described. Here, we identify MKV3 as a first-in-class inhibitor of Cu-transporting P-type ATPases that targets a conserved Cu+ entry site to the translocation pathway. In silico docking against the Xenopus ATP7B structure revealed a highly conserved pocket suitable for pharmacological inhibition. MKV3 bound human ATP7A and ATP7B with nanomolar affinity, competed with N-terminal metal-binding domains for access to the Cu+ entry site, and selectively inhibited Escherichia coli CopA ATPase activity and Cu+ transport. Mechanistically, MKV3 blocked chaperone-mediated Cu+ delivery to the intramembranous CPC site of CopA that is essential for its transport function. We further identified a single charged P-domain residue that governed MKV3 affinity and potency across species. Functionally, MKV3 phenocopied the genetic loss of Cu+-ATPases in bacteria, fungi, plants, zebrafish, and mammals, impairing copper-dependent enzymes, transporter trafficking, and copper tolerance. These findings establish a conserved, druggable vulnerability in Cu+-ATPases and introduce MKV3 as a broadly active chemical tool to modulate copper homeostasis across biological kingdoms. Significance StatementCopper-transporting P1B-type ATPases are essential for copper homeostasis in all domains of life, yet have lacked pharmacological inhibitors. This work identifies MKV3 as the first small-molecule inhibitor of Cu+-ATPases in bacteria, fungi, plants and animals, and defines a conserved, druggable Cu+ entry pocket that governs metal delivery to the transmembrane pathway. MKV3s ability to potentiate copper-mediated killing in multidrug-resistant bacterial pathogens highlights its potential as an antimicrobial adjuvant, while its attenuation of mammalian ATP7A/B function offers promise in oncology and copper-related diseases. Collectively, these findings establish a new tool for targeting of Cu+-ATPases with wide-ranging applications across biological systems.

cell biology↗

Extracellular vesicles facilitate the horizontal transfer of drug resistance and stem-like properties between ovarian tumor cells

Ovarian cancer stem cells (CSCs) can seed recurrent drug-resistant disease. Likewise, non-CSCs can acquire CSC phenotypic properties. How this process is orchestrated is of interest to inform how it might be prevented. We tested the hypothesis that ovarian CSC and/or drug-resistant tumor cells confer stem-like properties via extracellular vesicles (EVs). We focused our investigation on how EVs might mediate EZH2 signaling to promote a phenotypic change in drug-sensitive, non-CSCs. To accomplish this, we utilized paired PARP inhibitor-sensitive and - resistant ovarian cancer (OvCa) cell lines, EZH2 knockdown lines, and patient-derived organoids (PDOs) originating from recurrent high-grade serous OvCa. Small EVs isolated from drug-sensitive, CSC and/or drug-resistant enriched cultures, PARP inhibitor (olaparib) resistant lines, or drug-treated (olaparib or carboplatin) lines were cultured with treatment naive or sensitive lines for defined time points. The impact of small EV exposure was determined by assessing cell number, metabolic activity, viability, sphere and colony-forming capacity, ALDH activity, DNA damage, and changes in associated signaling pathways. We found that EVs from CSC or drug-resistant enriched cell fractions communicate CSC-like phenotypes to the more sensitive tumor cells via EZH2 canonical and non-canonical signaling pathways, promoting stemness. We conclude that EV-mediated activation of EZH2 signaling represents a targetable mechanism contributing to stemness-associated drug resistance in OvCa. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=160 SRC="FIGDIR/small/699925v1_ufig1.gif" ALT="Figure 1"> View larger version (44K): org.highwire.dtl.DTLVardef@6b4ffborg.highwire.dtl.DTLVardef@1501931org.highwire.dtl.DTLVardef@1a5f31aorg.highwire.dtl.DTLVardef@1fb475f_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

Mg2+-Dependent Multistep Folding and Stabilization of the GAAA Tetraloop-Receptor Interaction in a Group I Intron

Group I Introns are non-coding regions of pre-mRNA that catalyze their splicing from the RNA sequence by folding to a specific structure. We used computer simulations to study the folding mechanism of the P4-P6 domain in the Tetrahymena thermophila group I intron, focusing on the GAAA tetraloop-receptor (TL-R) interaction, which is a ubiquitous tertiary interaction in RNA structures. We show that the intron folds via a multistep pathway, populating seven states with distinct tertiary contacts. Under physiological Mg2+ concentrations ([Mg2+]), the loop-bulge-P4 tertiary interaction is essential to stabilize the docked TL-R complex, whereas in high [Mg2+], the TL-R complex is stable by itself. The solvated Mg2+ ions modulate the TL-R docking-undocking dynamics and stabilize non-native intermediate states. The condensation of Mg2+ in the major grooves of the TL and R helices is critical for them to attain specific stiffness essential for their facile docking. The results highlight the critical role of Mg2+ ions in facilitating TL-R interaction formation, which stabilizes long-range tertiary contacts in RNA structures. For Table of Contents Use Only O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=111 SRC="FIGDIR/small/700762v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@142852org.highwire.dtl.DTLVardef@1632ad1org.highwire.dtl.DTLVardef@190021aorg.highwire.dtl.DTLVardef@17a1261_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

Actin-dependent mechanotransduction controls nucleocytoplasmic partitioning of DNMT3a through ERK1/2 signaling during cutaneous wound healing

Cutaneous wound healing is a multifaceted physiological process that requires a cell state transition from homeostasis to tissue repair. An important contributor to this reprogramming is the wound-induced mechanical cues that are perceived by epidermal keratinocytes. Previously we identified that the nuclear translocation of the de novo DNA methyltransferase 3A (DNMT3a) upon wounding as an important regulator of this cell-state transition. However, the molecular mechanisms linking mechanotransduction to epigenetic regulation remain incompletely understood. Here we show that under homeostasis, active ERK1/2 phosphorylates DNMT3a, intramolecularly masking its nuclear localization signal (NLS), resulting in its cytoplasmic sequestration. Upon wounding actin cytoskeleton remodeling leads to the downregulation of the Extracellular Signal-Related Kinase 1/2 (ERK 1/2) pathway. Wound-induced ERK1/2 inactivation unmasks the NLS, enabling DNMT3a nuclear translocation. Collectively, these findings define a mechanotransduction-driven signaling axis linking cytoskeletal dynamics to epigenetic regulation and confirms an active role for differentiated keratinocytes in initiating early wound repair programs.

cell biology↗

Designing AI-programmable therapeutics with the EDEN family of foundation models

The ability to interpret, modify, and design DNA has driven many of the most significant advances in modern medicine, from diagnostics, biologics, and vaccines to cell and gene therapies. However, the inherent complexity of biological systems means that most modern medicines are still engineered using bespoke, labor-intensive processes. To address the need for a generalisable and programmable approach to therapeutic design, we introduce the EDEN (environmentally-derived evolutionary network) family of metagenomic foundation models, including a 28 billion parameter model trained on 9.7 trillion nucleotide tokens from BaseData1. This dataset, at the time of training, contained more than 10 billion novel genes from over 1 million new species, and is intentionally enriched for environmental and host-associated metagenomes, phage sequences, and mobile genetic elements, enabling the model to learn from diverse and novel cross-species evolutionary mechanisms and apply them to key challenges in human health. EDEN achieves state-of-the-art performance across a series of predictive and generative genomic and protein benchmarks. To demonstrate the models broad applicability across biology, we evaluate EDENs capacity for programmable therapeutic design by challenging a single architecture to design biological novelty across three distinct therapeutic modalities, disease areas and biological scales: (i) large gene insertion, (ii) antibiotic peptide design, and (iii) microbiome design. First, we demonstrate AI-programmable Gene Insertion (aiPGI), in which EDEN designs de novo large serine recombinases (LSRs) capable of inserting large pieces of DNA at desired target sites in the human genome when prompted only on 30 nucleotides of DNA sequence from the desired target site. In low-N experimental validation, EDEN generated multiple active recombinases for all tested disease-associated genomic loci (ATM, DMD, F9, FANCC, GALC, IDS, P4HA1, PHEX, RYR2, USH2A) and 4 potential safe harbor sites in the human genome. EDEN achieves an overall functional hit rate of 63.2% across diverse DNA prompts when prompted on only 30bp of DNA from outside the training data. 50% of EDEN-generated LSRs were active in human cells, achieving therapeutically relevant levels of CAR insertion in primary human T cells. We also show that EDEN can generate active bridge recombinases when prompted on the associated guide RNA alone, with sequence identities to training and public data as low as 65%. These results pave the way for a new generation of cell and gene therapies by opening the door to rapid, programmable and site-specific integration of large genetic payloads without double-strand breaks. This offers an alternative to the safety, efficiency and payload limitations inherent in viral or nuclease-based editing at thousands of currently intractable human therapeutic targets. Second, we use the same model to generate a focused low-N library of novel antimicrobial peptides where 97% showed activity, with top candidates achieving single-digit micromolar potency against critical-priority multidrug-resistant pathogens. Third, to demonstrate that EDEN captures inter-genomic features, we design a gigabase-scale microbiome with over 94,000 synthetic metagenomic assemblies, including prophage genomes and correct cross-species metabolic pathway completions. The EDEN-generated synthetic microbiome covers 9,067 species with a biome-specific taxonomic accuracy of 99%. Over 1,500 of the generated species were outside the fine-tuning dataset while retaining the correct microecological properties and biome association, thus significantly expanding genetic and taxonomic diversity. Together, these results establish a new strategic direction for AI-programmable therapeutics, in which a single foundation model architecture designs candidate therapeutics across diverse modalities and disease areas. This suggests that the combination of billions of years of evolutionary data with specific therapeutic records offers a clear, scaling-driven path to making therapeutic design a predictable engineering discipline. O_FIG O_LINKSMALLFIG WIDTH=141 HEIGHT=200 SRC="FIGDIR/small/699009v2_ufig1.gif" ALT="Figure 1"> View larger version (59K): org.highwire.dtl.DTLVardef@68c20borg.highwire.dtl.DTLVardef@19b8e8corg.highwire.dtl.DTLVardef@1ab9362org.highwire.dtl.DTLVardef@1592cb7_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

Liquid liquid phase separation of the intrinsically disordered protein JPT2compartmentalizes components of NAADP-evoked Ca2+ signaling

Nicotinic acid adenine dinucleotide phosphate (NAADP) is a Ca2+-releasing second messenger that activates two-pore channels (TPCs) on endosomes and lysosomes. Rather than binding TPCs directly, NAADP acts through cytoplasmic NAADP-binding proteins (NAADP-BPs) which are essential for endolysosomal Ca2+ release. Here we characterized the properties of two recombinant, purified NAADP-BPs: Jupiter Microtubule Associated Homolog 2 (JPT2) and like-Sm protein 12 (LSM12). In contrast to LSM12, JPT2 is predicted to be an intrinsically disordered protein, a feature confirmed by circular dichroism and NMR spectroscopy. Under conditions of low Na+ concentration or molecular crowding, JPT2 underwent phase separation, as demonstrated by multiple orthogonal approaches. JPT2 condensates displayed liquid-like behavior and efficiently recruited LSM12, a novel fluorescent NAADP analog, as well as tubulin. JPT2 condensates also interacted with polymerized microtubules and lysosomes isolated from human cell lines. These findings reveal an unexpected capability of NAADP-BPs to undergo phase separation, and segregate with components needed for NAADP-dependent Ca2+ release. We speculate that these signaling condensates dictate cellular NAADP sensitivity, desensitization of NAADP responses, as well as NAADP targeting to TPCs at membrane contact sites between acidic organelles and the endoplasmic reticulum. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=159 SRC="FIGDIR/small/698830v1_ufig1.gif" ALT="Figure 1"> View larger version (46K): org.highwire.dtl.DTLVardef@108ebc4org.highwire.dtl.DTLVardef@ae9eb9org.highwire.dtl.DTLVardef@36b859org.highwire.dtl.DTLVardef@81053a_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

circDiscoverer: A resource for curated and predicted circRNA-Protein interactions with experimental support tools

Circular RNAs (circRNAs) regulate various cellular processes by interacting with miRNAs, proteins, and by encoding peptides. Compared to circRNA-miRNA interactions, circRNA-protein interactions (CPIs) and circRNA modifications have been relatively less explored. Here, we developed a comprehensive user-friendly web resource, CircDiscoverer, to explore CPIs and modification landscapes across multiple species, including Homo sapiens, Mus musculus, Drosophila melanogaster, and Arabidopsis thaliana. CircDiscoverer integrates manually curated literature-supported CPIs with computationally predicted interactions. Furthermore, it offers comprehensive insights into protein-binding profiles and detailed information on interacting circRNAs, including qRT-PCR primer sequences, guide RNAs for experimental validation and potential RNA modifications, thereby providing an integrated platform for further research. CircDiscoverer is available at https://aclab.iiserb.ac.in.

bioinformatics↗

Efficacy of immunostimulant feed additive ShrimpGuard, for the preventive management of White Spot Syndrome Virus (WSSV) infection in shrimps

White spot syndrome virus (WSSV) is a virulent lethal viral pathogen affecting shrimp aquaculture worldwide. Current solutions for WSSV management can be divided into three categories; farm management, selective breeding or genetic engineering to get WSSV resistant varieties of shrimp, and developing vaccine or drug-like solutions specifically targeting WSSV. At present, farm management or good farming practice is the most reliable way of managing the problem, anti-WSSV vaccine, or anti-WSSV drugs that are reliable and scalable in the market are not available. We have developed a feed additive named "ShrimpGuard" using our SOLAQ (Solutions for Agriculture and Aquaculture) platform that specifically provides Shrimp protection against the WSSV infection. Here we report protection observed in both laboratory and farm demonstrations, in Penaeus vannamei. Collectively, these studies show that Shrimp-Guard does not cause any adverse effects on shrimp health and is able to provide significant protection to shrimp against WSSV infection. Briefly, the immune parameters of shrimp respond to ShrimpGuard treatment with increase observed in Total haemocyte count, and activity of prophenoloxidase. Further, survival rates of shrimp treated with ShrimpGuard after WSSV infection was between 80-100 % compared to 0% survival observed in untreated shrimp. The ShrimpGuards performance was tested in production farms and it was found that ShrimpGuard is safe for farm-based applications and the data obtained shows a positive impact on shrimp yield after ShrimpGuard treatment.

immunology↗

Highly Efficient Multiplexed Genome Engineering and Clone Selection to Enable Next Generation Induced Pluripotent Stem Cell (iPSC)-based Cell Therapies

Human induced pluripotent stem cells (iPSCs) have revolutionized regenerative medicine and cellular therapies. To improve the functionality and safety of iPSC-based therapies, genome engineering has been employed to disrupt gene expression and introduce therapeutic transgenes. However, current genome editing methods face significant challenges, including labor-intensive procedures, low efficiency, and safety concerns. Here, we report a novel and efficient approach for multiplex genome engineering in iPSCs using Integrase-mediated Programmable Genomic Integration (I-PGI) technology. I-PGI combines CRISPR-mediated genome editing and site-specific integrases, enabling precise gene insertion without DNA double-strand breaks. By optimizing I-PGI components and protocols, we achieved multiple gene knockouts and knock-ins of up to five transgenes in a single process. Additionally, we developed a streamlined workflow for enrichment, deposition, and screening of single-cell clones with the desired genome edits. This approach significantly accelerates and improves the precision of multiplex iPSC engineering, paving the way for next-generation iPSC-based therapies.

genomics↗

Multiplexed Pan Soluble Ligandome Assaying via OASIS

Screening soluble protein ligands is essential for understanding signaling interactions and enabling drug discovery. Currently, screens require arrayed formats because ligand diffusibility causes non-cell-autonomous effects. To enable multiplexed pooled assaying, we developed Obligate Autocrine Signaling In situ Screening (OASIS). Using lentiviral delivery of genetically barcoded ligands fused to a tethering domain, we anchor proteins to the expressing cells outer membrane, exclusively enforcing autocrine signaling. Validation using IFNA2 and EGF demonstrated uncompromised autocrine signaling with significantly reduced paracrine activity. We leveraged OASIS to perform a pan-ligandome fitness screen of all 770 validated human ligands in KOLF2.1J hiPSCs, identifying potent self-renewal factors, including FGF family ligands, which we experimentally validated in soluble form. Finally, we performed single-cell Perturb-Seq to map the transcriptional remodeling induced by the pan-ligandome library. OASIS provides a generalizable framework to accelerate functional interrogation of the human ligandome and novel peptide binders within live cells and diverse lineages. SUMMARYWe present Obligate Autocrine Signaling In situ Screening (OASIS), a platform enabling pooled assaying of soluble ligands by tethering genetically barcoded proteins to the surface of their expressing cells, enforcing autocrine signaling. We demonstrate OASIS by assaying all verified human ligands in hiPSCs, measuring fitness and transcriptional impacts. Our results recapitulate canonical interferon and FGF mediated signaling, validating the application of OASIS for rapid interrogation of ligands and engineered binders.

synthetic biology↗

Regulatable In Vivo Gene Expression via Adaptamers

Precise, reversible control of transgene expression is essential for safe and durable gene therapy, yet current inducible systems remain difficult to translate in vivo due to large size, limited induction duration, and dependence on immunogenic regulators. Here we present the adaptamer (ADAR modulatable aptamer), a compact (<120 bp) RNA switch that couples an FDA-approved small-molecule-responsive aptamer with endogenous ADAR-mediated RNA editing to regulate expression. Ligand binding stabilizes a double-stranded RNA structure that recruits ADAR to convert a stop codon into a sense codon, restoring downstream protein translation. This cleavage-free, post-transcriptional mechanism enables precise, small-molecule-dependent modulation without exogenous protein machinery. We demonstrate that adaptamers are highly functional across multiple cell lines, including human T-cells. In mice, AAV-delivered adaptamer-controlled FGF21 expression induced metabolic remodeling, significantly increasing energy expenditure and reversing obesity. This minimal, programmable system offers a clinically compatible approach for tunable genetic medicines and safe deployment of pleiotropic and dose-limited proteins.

synthetic biology↗

Small molecule inhibitor of PPARγ acetylation promotes insulin sensitization and browning of white adipose tissue with improved safety

The nuclear receptor PPAR{gamma} is a primary therapeutic target for insulin resistance and type 2 diabetes (T2D); however, its thiazolidinedione (TZD) class of PPAR{gamma} agonists have substantial safety concerns in clinical utilization. Genetic inhibition of PPAR{gamma} acetylation at K268 and K293 dissociates the major adverse effects of TZDs from insulin sensitization and other metabolic improvements. We therefore posit that chemical inhibition of PPAR{gamma} acetylation would elicit insulin sensitization with improved safety. Here we describe the identification of a synthetic thiopyrimidine derivative (TPMD) that acts as a small molecule inhibitor of PPAR{gamma} acetylation. TPMD improves insulin sensitivity, promotes brown remodeling of white adipose tissue (WAT), increases energy expenditure, and decreases adiposity in dietary and genetic mouse models of T2D. Importantly, TPMD is deprived of the major side effects of TZDs, including weight gain, fluid retention, cardiac hypertrophy, bone marrow adiposity, and bone loss. X-ray crystallography at 2.0 [A] resolution reveals a unique binding mode of TPMD molecules to the PPAR{gamma} ligand-binding domain and provides structural and molecular basis for the acetylation inhibitory activity of TPMD together with site-directed mutagenesis studies. These findings identify TPMD as a first-in-class compound that specifically targets PPAR{gamma} acetylation and possesses the potential of developing into a safe insulin sensitizer to treat and prevent T2D and obesity.

cell biology↗

Abiotic Stress-Mediated Modulation of the tncRNome: Unraveling tRNA-Derived RNA Networks in Plant Adaptive Responses

Transfer RNA-derived non-coding RNAs (tncRNAs) are potential regulators of gene expression in plants, and its crucial to understand their diversity and functional roles in modulating plant stress responses. Here, we have investigated abiotic stress-responsive tncRNAs in two important crops, rice and chickpea, under drought and salt stress conditions. Small RNA sequencing revealed that 70.2% of tncRNAs in rice and 45% in chickpea were commonly expressed across multiple stress conditions. qRT-PCR validation showed that most tRF-5s were upregulated in both rice and chickpea under drought and salt stress. In rice, 3'tRHs were specifically induced by drought, whereas 5'tRHs responded predominantly to salt stress. In chickpea, tRF-3s were upregulated during drought but downregulated under salt stress. Furthermore, we confirmed the mRNA targets of these stress-associated tRNA fragments using RLM-RACE, thereby verifying the cleavage of specific target mRNAs in both species. Expression profiling of target genes revealed a negative association with their corresponding tRFs in both species. Subcellular localization demonstrated that OsGEM and OsPPR were primarily localized in the nucleus, while CaMIZ1 and CaSKP1 were detected at the plasma membrane. Overall, we identified abiotic stress-responsive tncRNAs from rice and chickpea that suggest their potential contribution to plant resilience under changing environmental conditions.

bioinformatics↗