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Zhao, J.

Publications and source records attributed to Zhao, J..

At least 235 records · Page 13Linked to original sources

scMODAL: A general deep learning framework for comprehensive single-cell multi-omics data alignment with feature links

Recent advancements in single-cell technologies have enabled comprehensive characterization of cellular states through transcriptomic, epigenomic, and proteomic profiling at single-cell resolution. These technologies have significantly deepened our understanding of cell functions and disease mechanisms from various omics perspectives. As these technologies evolve rapidly and data resources expand, there is a growing need for computational methods that can integrate information from different modalities to facilitate joint analysis of single-cell multi-omics data. However, integrating single-cell omics datasets presents unique challenges due to varied feature correlations and technology-specific limitations. To address these challenges, we introduce scMODAL, a deep learning framework tailored for single-cell multi-omics data alignment using feature links. scMODAL integrates datasets with limited known positively correlated features, leveraging neural networks and generative adversarial networks to align cell embeddings and preserve feature topology. Our experiments demonstrate scMODALs effectiveness in removing unwanted variation, preserving biological information, and accurately identifying cell subpopulations across diverse datasets. scMODAL not only advances integration tasks but also supports downstream analyses such as feature imputation and feature relationship inference, offering a robust solution for advancing single-cell multi-omics research.

bioinformatics↗

Iterative sacrificial 3D printing and polymer casting to create complex vascular grafts and multi-compartment bioartificial organs

Several emerging strategies to engineer artificial organs employ 3D printing to create vascular templates to provide nutrients and oxygen to immobilized cells. Significant challenges emerge when considering clinical implementation such as immune rejection of allogeneic cell sources, as well as achieving adequate perfusion and integration with endogenous vasculature. We propose a method by which cell-laden hydrogels are molded around ready-made polymeric vascular templates created via 3D printing to create human-scale artificial organs with internal vasculature. We applied this technique to create bioartificial pancreas systems with up to 9 internal flow channels via sacrificial carbohydrate glass 3D printing, porogen-loaded polycarbonate polyurethane dip-coating, followed by casting cell-laden hydrogels around the vascular templates. We optimized porogen size and concentration to maximise the porosity of our scaffolds without compromising mechanical properties, resulting in suture retention strength and compliance respectively matching commercial vascular grafts and native vessels. Bioreactor perfusion studies showed survival and maturation of stem cell derived pancreatic islets without significant differences to traditional suspension culture protocols. Insulin response dynamics were rapid in response to a glucose challenge at the perfusion inlet. Transplantation of the devices as iliac arteriovenous shunts in nondiabetic pigs confirmed safety and patency. These results show promise for the development of an implantable vascularized pancreas for the treatment of type 1 diabetes and demonstrate how bioartificial organs with engineered vascular geometries can be designed for translational applications.

bioengineering↗

INSPIRE: interpretable, flexible and spatially-aware integration of multiple spatial transcriptomics datasets from diverse sources

Recent advances in spatial transcriptomics technologies have led to a growing number of diverse datasets, offering unprecedented opportunities to explore tissue organizations and functions within spatial contexts. However, it remains a significant challenge to effectively integrate and interpret these data, often originating from different samples, technologies, and developmental stages. In this paper, we present INSPIRE, a deep learning method for integrative analyses of multiple spatial transcriptomics datasets to address this challenge. With designs of graph neural networks and an adversarial learning mechanism, INSPIRE enables spatially informed and adaptable integration of data from varying sources. By incorporating non-negative matrix factorization, INSPIRE uncovers interpretable spatial factors with corresponding gene programs, revealing tissue architectures, cell type distributions and biological processes. We demonstrate the capabilities of INSPIRE by applying it to human cortex slices from different samples, mouse brain slices with complementary views, mouse hippocampus and embryo slices generated through different technologies, and spatiotemporal organogenesis atlases containing half a million spatial spots. INSPIRE shows superior performance in identifying detailed biological signals, effectively borrowing information across distinct profiling technologies, and elucidating dynamical changes during embryonic development. Furthermore, we utilize INSPIRE to build 3D models of tissues and whole organisms from multiple slices, demonstrating its power and versatility.

bioinformatics↗

MAPK/ERK signaling in gliomas modulates interferon responses, T cell recruitment, enhances tumor-microglia crosstalk, and drives immune checkpoint blockade efficacy.

BackgroundGlioblastoma (GB) remains a formidable challenge in neuro-oncology, with immune checkpoint blockade (ICB) showing limited efficacy in unselected patients. We previously recently established that MAPK/ERK signaling is associated with overall survival following anti-PD-1 and anti-CTLA-4 treatment in recurrent GB. However, the causal relationship between MAPK/ERK signaling and susceptibility to ICB, as well as the mechanisms underlying this association, remain poorly understood. MethodWe conducted in vivo kinome-wide CRISPR/Cas9 screenings in murine gliomas to identify key regulators of susceptibility to anti-PD-1 and CD8+ T cell responses and performed survival studies to validate the most relevant genes. Additionally, paired single cell RNA- sequencing (scRNA-seq) with p-ERK staining, spatial transcriptomics on GB samples, and ex-vivo slice culture of a BRAFV600E mutant GB tumor treated with BRAFi/MEKi were used to determine the causal relationship between MAPK signaling, tumor cell immunogenicity, and modulation of microglia phenotype. ResultsCRISPR/Cas9 screens identified the MAPK pathway, particularly the RAF-MEK-ERK pathway, as the most critical modulator of glioma susceptibility to CD8+ T cells, and anti-PD-1 across all kinases. Experimentally-induced ERK phosphorylation in gliomas enhanced survival with ICB treatment, led to durable anti-tumoral immunity upon re-challenge and memory T cell infiltration in long-term survivors. Elevated p-ERK in glioma cells correlated with increased interferon responses, antigen presentation and T cell infiltration in GB. Moreover, spatial transcriptomics and scRNA-seq analysis revealed the modulation of interferon responses by the MAPK/ERK pathway in BRAFV600E human GB cells with ERK1/2 knockout and in slice cultures of human BRAFV600E GB tissue. Notably, BRAFi/MEKi treatment disrupted the interaction between tumor cells and tumor-associated macrophages/microglia in slice cultures from BRAFV600E mutant GB. ConclusionOur data indicate that the MAPK/ERK pathway is a critical regulator of GB cell susceptibility to anti-tumoral immunity, modulating interferon responses, and antigen-presentation in glioma cells, as well as tumor cell interaction with microglia. These findings not only elucidate the mechanistic underpinnings of immunotherapy resistance in GB but also highlight the MAPK/ERK pathway as a promising target for enhancing immunotherapeutic efficacy in this challenging malignancy.

cancer biology↗

Distinct roles of general anesthesia activated CeA neurons in acute versus late phase of neuropathic pain

A previous study discovered a distinct population of GABAergic neurons in the central amygdala (CeA) that can be activated by general anesthesia (CeAGA) and exert analgesic functions (Hua et al., 2020). To independently reproduce these prior findings and to investigate the electrophysiological properties of CeAGA neurons, we first used 1.2% isoflurane to induce c-Fos activation in the mouse brain and validated the Fos expression by RNAscope in situ hybridization. Indeed, isoflurane induced robust Fos expression in CeA and these Fos+ CeAGA neurons are GABAergic neurons (Vgat+). We next used Fos-TRAP2 method (different from the CANE method used in the prior study) to label CeAGA neurons (tdTomato+). Our ex vivo electrophysiological recordings in brain slices revealed that compared to Fos-negative CeA neurons, CeAGA neurons had significantly higher excitability and exhibited distinct patterns of action potentials. Chemogenetic activation of Fos-TRAPed CeAGA neurons was effective at increasing pain thresholds in naive mice and mice with early-phase neuropathic pain 2 weeks after spared nerve injury (SNI). However, the same chemogenetic activation of CeAGA neurons only had modest analgesia in the late phase of SNI at 8 weeks, although it was highly effective in reducing chronic pain-associated anxiety behaviors at this stage. We found that Fos-negative CeA neurons, but not CeAGA neurons, exhibited increased excitability in the late-phase of SNI, suggesting that chronic pain causes a shift in the relative activity of the CeA microcircuit. Interestingly, Fos-negative neurons exhibited much higher expression of K+-Cl- cotransporter-2 (KCC2), and KCC2 expression was downregulated in the CeA in the late-phase of neuropathic pain. These results support the idea that targeting CeAGA neurons may provide therapeutic benefits for pain relief and chronic pain-associated anxiety. Our findings also suggest distinct roles of CeAGA neurons in regulating physiological pain, acute pain, and chronic pain with a possible involvement of KCC2.

neuroscience↗

Development of a DNA-encoded library screening method DEL Zipper to empower the study of RNA-targeted chemical matter

To date, RNA-targeted chemical matter is under explored due to a lack of robust screening assays. In this study, we present a novel RNA-targeted small molecule screening approach using a specialized DNA-encoded library (DEL). Our findings reveal that the specialized DEL library, called "DEL Zipper", can significantly reduce single-stranded DNA-RNA region interaction signals during various kinds of RNA selection. By performing the selection against both G-quadruplex, we have identified novel hits that interact with RNA targets and the results are validated through binding. This study demonstrates that the "DEL Zipper" method is a robust screening assay that has potential for discovering small molecule ligands for diverse RNA targets.

molecular biology↗

Direct detection of endogenous Gαi activity in cells with a sensitive conformational biosensor

Activation of heterotrimeric G-proteins (G{beta}{gamma}) by G-protein-coupled receptors (GPCRs) is not only a mechanism broadly used by eukaryotes to transduce signals across the plasma membrane, but also the target for a large fraction of clinical drugs. However, approaches typically used to assess this signaling mechanism by directly measuring G-protein activity, like optical biosensors, suffer from limitations. On one hand, many of these biosensors require expression of exogenous GPCRs and/or G-proteins, compromising readout fidelity. On the other hand, biosensors that measure endogenous signaling may still interfere with the signaling process under investigation or suffer from having a small dynamic range of detection, hindering broad applicability. Here, we developed an optical biosensor that detects the endogenous G-protein active species Gi-GTP upon stimulation of endogenous GPCRs more robustly than current state-of-the-art sensors for the same purpose. Its design is based on the principle of bystander Bioluminescence Resonance Energy Transfer (BRET) and leverages the Gi-binding protein named GINIP as a high affinity and specific detector module of the GTP-bound conformation of Gi. We optimized this design to prevent interference with Gi-dependent signaling (cAMP inhibition) and to enable implementation in different experimental systems with endogenous GPCRs, including neurotransmitter receptors in primary astroglial cells or opioid receptors in cell lines, which revealed opioid neuropeptide-mediated activation profiles different from those observed with other biosensors involving exogenous GPCRs and G-proteins. Overall, we introduce a biosensor that directly and sensitively detects endogenous activation of G-proteins by GPCRs across different experimental settings without interfering with the subsequent propagation of signaling.

pharmacology and toxicology↗

Seq2Topt: a sequence-based deep learning predictor of enzyme optimal temperature

An accurate deep learning predictor is needed for enzyme optimal temperature (Topt), which quantitatively describes how temperature affects the enzyme catalytic activity. Seq2Topt, developed in this study, reached a superior accuracy on Topt prediction just using protein sequences (RMSE = 13.3 and R2=0.48) in comparison with existing models, and could capture key protein regions for enzyme Topt with multi-head attention on residues. Through case studies on thermophilic enzyme selection and predicting enzyme Topt shifts caused by point mutations, Seq2Topt was demonstrated as a promising computational tool for enzyme mining and in-silico enzyme design. Additionally, accurate deep learning predictors of enzyme optimal pH (Seq2pHopt, RMSE=0.92 and R2=0.37) and melting temperature (Seq2Tm, RMSE=7.57 and R2=0.64) were developed based on the model architecture of Seq2Topt, suggesting that the development of Seq2Topt could potentially give rise to a useful prediction platform of enzymes.

bioinformatics↗

Therapeutic targeting of Syndecan-1 axis overcomes acquired resistance to KRAS-targeted therapy in gastrointestinal cancers

The therapeutic benefit of recently developed mutant KRAS (mKRAS) inhibitors has been limited by the rapid onset of resistance. Here, we aimed to delineate the mechanisms underlying acquired resistance to mKRAS inhibition and identify actionable targets for overcoming this clinical challenge. Previously, we identified Syndecan-1 (SDC1) as a key effector for pancreatic cancer progression whose surface expression is driven by mKRAS. By leveraging both pancreatic and colorectal cancer models, we found that surface SDC1 expression was initially diminished upon mKRAS inhibition, but recovered in tumor cells that bypass mKRAS dependency. Functional studies showed that these tumors depended on SDC1 for survival, further establishing SDC1 as a driver for the acquired resistance to mKRAS inhibition. Mechanistically, we revealed that the YAP1-SDC1 axis was the major driving force for bypassing mKRAS dependency to sustain nutrient salvage machinery and tumor maintenance. Specifically, YAP1 activation mediated the recovery of SDC1 localization on cell surface that sustained macropinocytosis and enhanced the activation of multiple RTKs, promoting resistance to KRAS-targeted therapy. Overall, our study has provided the rationale for targeting the YAP-SDC1 axis to overcome resistance to mKRAS inhibition, thereby revealing new therapeutic opportunities for improving the clinical outcome of patients with KRAS-mutated cancers.

cancer biology↗

Structural basis of human 20S proteasome biogenesis

New proteasomes are produced to accommodate increases in cellular catabolic demand and prevent the accumulation of cytotoxic proteins. Formation of the proteasomal 20S core complex relies on the function of the five chaperones PAC1-4 and POMP. To understand how these chaperones facilitate proteasome assembly, we tagged the endogenous chaperones using CRISPR/Cas gene editing and examined the chaperone-bound complexes by cryo-EM. We observed an early -ring intermediate subcomplex that is stabilized by PAC1-4, which transitions to {beta}-ring assembly upon dissociation of PAC3/PAC4 and rearrangement of the PAC1 N-terminal tail. Completion of the {beta}-ring and dimerization of half-proteasomes repositions critical lysine K33 to trigger cleavage of the {beta} pro-peptides, leading to the concerted dissociation of POMP and PAC1/PAC2 to yield mature 20S proteasomes. This study reveals structural insights into critical points along the assembly pathway of the human proteasome and provides a molecular blueprint for 20S biogenesis.

biochemistry↗

Ganoderma lucidum alleviates kidney aging by modulating gut microbiota

BackgroundGanoderma lucidum, a renowned traditional Chinese medicine known for its multiple health benefits, including anti-aging effect, has the ability to influence gut microbiota. However, its specific role in alleviating aging via gut microbiota has not been thoroughly investigated. Here, we explored the impact of G. lucidum sporoderm-broken spore powder (Gl-SBSP) on kidney aging using naturally aged and radiation-induced premature senescence mouse models. MethodsWe analyzed gut microbiota through high-throughput sequencing in naturally aged and radiation-induced premature senescence mice after oral administration of Gl-SBSP. Additionally, untargeted metabolomics of the gut microbiome in naturally aged mice was performed using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) analyses. Transcriptomics was employed to systematically measure gene expression levels throughout kidney tissues. ResultsGl-SBSP effectively mitigated aging-related phenotypic changes and significantly alleviated kidney aging, although outcomes were mixed in the irradiation mouse model. Importantly, Gl-SBSP modulated gut bacterial composition, particularly enriching Lachnospiraceae in naturally aged mice and in radiation-induced premature senescence mice with positive prognostic outcomes. Nicotinamide riboside (NR), derived from Lachnospiraceae bacteria, was identified as a potential metabolite that promoted recovery of senescent kidneys and improved renal function, validated through in vitro and in vivo experiments. Transcriptome analysis revealed that NR regulates kidney steroid metabolism in aging. ConclusionOur findings suggest that Gl-SBSPs anti-aging effect is mediated by NR derived from the Lachnospiraceae family, influencing kidney steroid metabolism via the gut-kidney axis. These results provide new insights into the mechanisms underlying the anti-aging properties of Gl-SBSP. AbstractGanoderma lucidum, also known as "Linzhi," is a well-established traditional Chinese medicine renowned for its multiple health benefits and its influence on gut microbiota. In this study, we investigated the anti-aging effect of G. lucidum sporoderm-broken spore powder (Gl-SBSP) through the gut-kidney axis using naturally aged and radiation-induced premature senescence mouse models. Administration of Gl-SBSP not only effectively mitigated aging-related phenotypic changes but also significantly alleviated kidney aging, although outcomes were mixed in the irradiation model. Importantly, Gl-SBSP modulated gut bacterial composition, particularly enriching Lachnospiraceae in naturally aged mice and in radiation-induced premature senescence mice with favorable and long-lasting outcomes. Nicotinamide riboside (NR), derived from Lachnospiraceae bacteria, was identified as a potential metabolite that facilitated the rejuvenation of senescent kidneys and improved renal function through both in vitro and in vivo experiments. Further investigation revealed that NR regulated steroid metabolic process in kidney. Our findings suggest that the anti-aging effect of Gl-SBSP is mediated by NR derived from the Lachnospiraceae bacteria, thereby influencing kidney steroid metabolism through the gut-kidney axis. This study provides novel insights into the anti-aging mechanisms of Ganoderma.

microbiology↗

Genetic variation in the species Arabidopsis thaliana reveals the existence of natural heat resilience factors for meiosis

Heat interferes with multiple meiotic processes leading to genome instability and sterility in flowering plants, including many crops. Despite its importance for food security, the mechanisms underlying heat tolerance of meiosis are poorly understood. In this study, we analyzed different meiotic processes in the Arabidopsis (Arabidopsis thaliana) accessions Columbia (Col) and Landsberg erecta (Ler), their F1 hybrids and F2 offspring under heat stress (37{degrees}C). At 37{degrees}C, Col exhibits significantly reduced formation of double-stand breaks (DSBs) and completely abolished homolog pairing, synapsis and crossover (CO) formation. Strikingly, Ler and Ler/Col hybrids are much less affected than Col. Interestingly, only 10% [~] 20% of F2 offspring exhibit the same heat tolerance of meiotic recombination as parents, indicating that heat resilience in Ler is controlled by the interplay of several loci. Moreover, F2 offspring show defective chromosome condensation in interkinesis, and untimely sister-chromatid segregation and/or chromosome fragmentation, the levels of which exceed those in either inbreds and/or hybrids thus implying a transgressive effect on heat tolerance of meiosis. Furthermore, correlation and cytogenetic analysis suggest that homolog pairing and/or synapsis have an impact on heat tolerance of chromosome morphology and stability during post-recombination stages under heat stress. Taken together, this study reveals the existence of natural heat resilience factors for meiosis in Arabidopsis, which have the great potential to be exploited in breeding programs. Author summaryEnvironmental temperature alterations affect meiotic recombination and/or chromosome segregation thus perturbing genetic makeup and genome stability in plants. We have previously reported that CO formation is fully abolished in Arabidopsis thaliana accession Col under heat stress (36{degrees}C-38{degrees}C) due to reduced DSB formation and impaired homolog pairing. Here, we show that in Arabidopsis thaliana accession Ler under the same high temperature conditions, both DSB and CO formation occur normally, and homolog pairing is mildly impacted, which indicate a striking difference in heat tolerance of meiotic recombination from Col. Remarkably, Col/Ler hybrids display the same heat tolerance as Ler, however, only 10% [~] 20% of F2 offspring behave the same as parents. Moreover, we found higher levels of defects in chromosome morphology and integrity, and sister-chromatid segregation in F2 population than those in both inbreds and hybrids, which suggest a transgressive effect influencing heat tolerance of meiosis. Our findings reveal that heat resilience in Arabidopsis is controlled by the interplay of multiple genomic loci, holding a great potential to be exploited in crop breeding.

plant biology↗

Loss of PINK1 causes age-dependent mitochondrial trafficking deficits in nigrostriatal dopaminergic neurons through aberrant activation of p38 MAPK

Mutations in PTEN-induced putative kinase 1 (PINK1) cause early-onset, autosomal-recessive Parkinsons disease (PD). While previous studies showed age-related declines in dopamine release and ATP levels in Pink1-/- mice, the mechanisms remain unclear. Using a novel TH-Mito-Dendra2 transgenic mouse model to label dopaminergic neuron mitochondria, it is found that PINK1 loss leads to age-dependent deficits in mitochondrial axonal trafficking, characterized by reduced anterograde movement and increased static mitochondria in acute brain slices, which more closely mimic in vivo conditions. Pharmacological induction of reactive oxygen species (ROS) and calcium release impaired mitochondrial mobility. Pink1 knockout mice exhibited elevated mitochondrial calcium, ROS levels, and p38 MAPK hyperactivation. Treatment with p38 inhibitor SB202190 restored mitochondrial motility and increased anterograde transport. Our findings suggest that PINK1 loss disrupts mitochondrial trafficking by disturbing calcium homeostasis and ROS production via the p38 pathway, contributing to PD pathogenesis. Table of Contents O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=181 SRC="FIGDIR/small/602426v3_ufig1.gif" ALT="Figure 1"> View larger version (68K): org.highwire.dtl.DTLVardef@1352742org.highwire.dtl.DTLVardef@a48086org.highwire.dtl.DTLVardef@1f209daorg.highwire.dtl.DTLVardef@164ed23_HPS_FORMAT_FIGEXP M_FIG C_FIG Using a novel mouse model with labeled dopaminergic neuron mitochondria, Zhao J. et al. discovered age dependent disruptions in mitochondrial transport. Their research showed progressive deterioration of mitochondrial movement, characterized by reduced anterograde movement and increased static mitochondria. These changes were associated with higher oxidative stress and hyperactivation of p38 MAPK in Parkinsons disease.

neuroscience↗

A new yeast-based bioreporter for simple, sensitive, and cost-effective detection of dioxin-like compounds

Dioxin-like compounds (DLCs) are environmental xenobiotics that can activate the aryl hydrocarbon receptors (AhR), thereby imposing a significant threat to human health through biomagnifications processes. In this study, a dioxin-activated nano-luminescent Saccharomyces cerevisiae bioreporter, called DnaSc, was developed for simple and rapid detection of DLCs and AhR agonists. The bioreporter used nano-luciferase NLuc as a signal generator to emit bioluminescent signals in response to DLCs without cell lysis. Through optimizing ARNT expression and engineering the aryl hydrocarbon receptor (AhR), the yeast-based bioassay exhibited a detection limit of 10 fM for 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) within 6 h, making it the most sensitive whole-cell biosensor reported to date. Furthermore, the detection capacity of the DnaSc bioassay for DLCs and AhR agonists was characterized. In summary, the yeast-based bioreporter developed in this study provided a simple, sensitive, and cost-effective method for DLCs detection.

bioengineering↗

Divergent roles of SOX2 in human and mouse germ cell specification related to X-linked gene dosage effects

Human primordial germ cell-like cells (hPGCLCs) can be generated from pluripotent stem cells (PSCs) but the differentiation efficiency of female hPSCs is often lower than that of male hPSCs. Moreover, Klinefelter Syndrome (KS), a condition characterized by an extra X-chromosome in males, often presents the failure of germline specification and infertility. In this study, we investigate how X-linked gene dosage affects hPGCLCs specification potential in both healthy and diseased conditions. We reveal that the X-chromosome plays a multifaceted role in modulating hPGCLCs induction. The inhibitory effects on TGF-beta/Activin A and BMP pathways by escape genes IGSF1 and CHRDL1, respectively, are demonstrated by the increased yield of hPGCLCs with knockdown experiments. Importantly, our results identified the intriguing role of SOX2 that is upregulated by the escape gene USP9X in hPGCLCs specification, highlighting a species-specific difference from the mouse model. The elevated USP9X-SOX2 regulatory axis profoundly influences cellular metabolism, mitochondrial morphology, and progenitor competence, thereby affecting hPGCLCs induction. Furthermore, the inability to downregulate SOX2 and upregulate SOX17 in response to BMP signaling impedes downstream gene activation due to motif binding competition. These findings shed novel insights into the hPGC specification by elucidating the differential roles of SOX2 versus SOX17 between mice and humans, influenced by X-linked gene dosage effects. Additionally, our results offer potential applications for improving the induction and survival efficiency of hPGCLCs from hPSCs, facilitating disease modeling and mechanistic studies. HighlightsO_LIDownregulation of three X-linked genes, i.e. IGSF1, CHRDL1 and USP9X, enhanced the differentiation efficiency of hPGCLCs C_LIO_LISOX2 as a downstream of human-specific escape gene USP9X plays a multifacet role against hPGCLCs specification C_LIO_LIFailure to timely downregulate SOX2 and upregulate SOX17 interferes downstream gene activation likely due to motif binding competition C_LI

cell biology↗

Dissecting the genetic regulation of lateral root development in tomato under salt stress

Roots are the primary site for sensing salt, making them critical to understanding how plants adapt to salinity. Unraveling the genetic and molecular mechanisms that maintain root growth under salt stress is essential for developing resilient crops. Tomatoes are vulnerable to salinity, as over 50% of arable land is projected to become saline by 2050. While salt stress effects on tomato seed germination, shoot growth, and fruit yield are well-documented, the genetic basis of root development under salinity remains underexplored. Previous studies focused on physiological responses of roots in a narrow range of cultivated-tomatoes, overlooking the genetic diversity for salt resilience in wild-relatives like Solanum pimpinellifolium. Here, we investigated salt-induced changes in root system architecture (RSA) across a natural diversity panel of 220 wild- and 25 cultivated-tomato varieties. We identified tolerant accessions with different RSA strategies; prioritizing lateral root elongation versus emergence. To identify genes involved in specific aspects of lateral root development, an F1 hybrid was generated that exhibited both parents combined characteristics. An F2-segregating population was then used to identify four distinct subpopulations for Bulk Segregant Analysis (BSA). Simultaneously, we conducted Genome-Wide-Association Studies (GWAS) on root architecture of wild-tomato accessions under salt stress. By integrating the results of BSA and GWAS, we identified 22 candidates involved in the maintenance of root architecture. We utilized RNA-Seq to examine transcriptome reprogramming of the 22 genes in tomato accessions with contrasting lateral root responses. This approach resulted in the identification of 2 genes, AP2-like-ethylene-responsive transcription factor TOE3 and L-ascorbate peroxidase. Using exogenous ethylene, we demonstrated that root architecture exhibits plasticity, which benefits the plant by reducing Na+ accumulation. We profiled H2O2 waves across root systems of tomatoes, emphasizing the role of ROS in maintaining root system architecture under salt stress. These findings provide novel genetic targets for enhancing salt resilience in tomatoes, opening avenues for future research and breeding programs.

plant biology↗

Structural Basis for Substrate Binding, Catalysis and Inhibition of Breast Cancer Target Mitochondrial Creatine Kinase by Covalent Inhibitor via Cryo-EM

Mitochondrial creatine kinases are key players in maintaining energy homeostasis in cells by working in conjunction with cytosolic creatine kinases for energy transport from mitochondria to cytoplasm. High levels of MtCK observed in Her2+ breast cancer and inhibition of breast cancer cell growth by substrate analog, cyclocreatine, indicate dependence of cancer cells on the energy shuttle for cell growth and survival. Hence, understanding the key mechanistic features of creatine kinases and their inhibition plays an important role in the development of cancer therapeutics. Herein, we present the mutational and structural investigation on understudied ubiquitous mitochondrial creatine kinase (uMtCK). Our cryo-EM structures and biochemical data on uMtCK showed closure of the loop comprising residue His61 is specific to and relies on creatine binding and the reaction mechanism of phosphoryl transfer depends on electrostatics in the active site. In addition, the previously identified covalent inhibitor CKi showed inhibition in breast cancer BT474 cells, however our biochemical and structural data indicated that CKi is not a potent inhibitor for breast cancer due to strong dependency on the covalent link formation and inability to induce conformational changes upon binding.

biochemistry↗

Inhibitor of cardiolipin biosynthesis-related enzyme MoGep4 confers broad-spectrum anti-fungal activity

Plant pathogens cause devastating diseases, leading to serious losses to agriculture. Mechanistic understanding of pathogenesis of plant pathogens lays the foundation for the development of fungicides for disease control. Mitophagy, a specific form of autophagy, is important for fungal virulence. The role of cardiolipin, mitochondrial signature phospholipid, in mitophagy and pathogenesis is largely unknown in plant pathogenic fungi. The functions of enzymes involved in cardiolipin biosynthesis and relevant inhibitors were assessed using a set of assays, including genetic deletion, plant infection, lipidomics, chemical-protein interaction, chemical inhibition, and field trials. Our results showed that the cardiolipin biosynthesis-related gene MoGEP4 of the rice blast fungus Magnaporthe oryzae regulates growth, conidiation, cardiolipin biosynthesis, and virulence. Mechanistically, MoGep4 regulated mitophagy and Mps1-MAPK phosphorylation, which are required for virulence. Chemical alexidine dihydrochloride (AXD) inhibited the enzyme activity of MoGep4, cardiolipin biosynthesis and mitophagy. Importantly, AXD efficiently inhibited the growth of 10 plant pathogens and controlled rice blast and Fusarium head blight in the field. Our study demonstrated that MoGep4 regulates mitophagy, Mps1 phosphorylation and pathogenesis in M. oryzae. In addition, we found that the MoGep4 inhibitor, AXD, displays broad-spectrum antifungal activity and is a promising candidate for fungicide development.

microbiology↗