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PubMed2026

Beyond Earth: Recent Advancements in Microgravity Biomedical and Genetic Research in Saudi Arabia.

Microgravity research has emerged as a rapidly evolving field at the intersection of space medicine, genomics, biotechnology, and precision medicine. Exposure to the space environment induces complex physiological and molecular adaptations that affect multiple biological systems, including immune regulation, metabolism, musculoskeletal function, and gene expression. Recent advances in genomics, multi-omics technologies, artificial intelligence, and bioengineering have substantially improved our understanding of biological adaptation to spaceflight and expanded opportunities for translational biomedical research. This review summarizes recent advances in genetic and biomedical research under microgravity conditions, with particular emphasis on molecular mechanisms, omics technologies, genome editing, microbiome research, regenerative medicine, and personalized healthcare approaches. Major experimental platforms, landmark spaceflight studies, and translational applications in infectious diseases, cancer biology, aging, tissue engineering, and pharmaceutical development are discussed. The review also highlights Saudi Arabia's emerging contributions to genomic medicine and space biosciences through initiatives such as the Saudi Human Genome Program, the Saudi Pangenome Project, the Saudi Space Agency, and the BioGravity Initiative. Recent Saudi participation in human spaceflight and microgravity-associated biomedical research is discussed within the context of Vision 2030 and national investments in biotechnology and precision medicine. Collectively, advances in microgravity research are expected to contribute to the advancement of precision medicine and facilitate the development of innovative diagnostic and therapeutic strategies with significant implications for both human space exploration and terrestrial healthcare.

Humans↗
PubMed2025

Integrated epigenetic and genetic programming of primary human T cells.

Targeted epigenetic engineering of gene expression in cell therapies would allow programming of desirable phenotypes without many of the challenges and safety risks associated with double-strand break-based genetic editing approaches. Here, we develop an all-RNA platform for efficient, durable and multiplexed epigenetic programming in primary human T cells, stably turning endogenous genes off or on using CRISPRoff and CRISPRon epigenetic editors. We achieve epigenetic programming of diverse targeted genomic elements without the need for sustained expression of CRISPR systems. CRISPRoff-mediated gene silencing is maintained through numerous cell divisions, T cell stimulations and in vivo adoptive transfer, avoiding cytotoxicity or chromosomal abnormalities inherent to multiplexed Cas9-mediated genome editing. Lastly, we successfully combined genetic and epigenetic engineering using orthogonal CRISPR Cas12a-dCas9 systems for targeted chimeric antigen receptor (CAR) knock-in and CRISPRoff silencing of therapeutically relevant genes to improve preclinical CAR-T cell-mediated in vivo tumor control and survival.

Humans↗
PubMed2026

Evaluation of germline transmission of electroporation-mediated double gene-edited cattle lines.

Gene editing in livestock using clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) offers a promising approach for genetic improvement in cattle. This study evaluated germline transmission and mutation stability of double-knockout cattle generated by zygote electroporation. Previously reported myostatin/beta-lactoglobulin (MSTN/BLG) and newly generated α-1,3-galactosyltransferase (GGTA1/BLG) double-knockout cattle were produced using CRISPR/Cas9-mediated genome editing. Targeted deep sequencing demonstrated extensive somatic mosaicism across multiple tissues. Computer-assisted sperm analysis (CASA) demonstrated normal sperm motility in MSTN/BLG double-knockout males. Fertilization of wild-type oocytes produced heterozygous embryos, with mutation frequencies of 37.76 ± 10.74% at the MSTN locus and 54.80 ± 7.73% at the BLG locus, as assessed by T7 endonuclease I (T7E1) assay. MSTN/BLG double-knockout sperm were subsequently used for embryo production and for artificial insemination of GGTA1/BLG double-knockout females. Healthy offspring were successfully obtained (n = 3), alongside one stillborn calf. Targeted deep sequencing of all four progenies revealed highly variable allele frequencies that deviated substantially from the approximately 50% expected for heterozygous germline transmission. In contrast, whole-genome sequencing (WGS) results were consistent with Mendelian expectations, underscoring the limitations of PCR-based targeted sequencing for assessing germline transmission in mosaic founders. These results show that CRISPR/Cas9-edited embryos generated by electroporation can develop into healthy, sexually mature cattle capable of germline transmission. While variable transmission rates were observed owing to founder mosaicism, non-mosaic F1 offspring were successfully generated. However, direct, embryo-mediated gene-editing strategies remain technically and economically challenging for large-scale commercial calf production, and reports in cattle are limited. This study provides a reference for future applications of gene-edited embryos and their germline propagation.

Animals↗
PubMed2026

Protocol for robust gene knockout and reliable validation in human cell lines using quad-guide RNA vectors.

CRISPR-Cas9 is a powerful tool for editing genomic loci, however achieving high knockout efficiency at certain targets remains challenging. Here, we present a protocol for gene knockout using an all-in-one, quad-guide RNA-expressing vector. We describe steps for plasmid construction, virus preparation, transduction, and subsequent gene editing and functional validation within DLD-1 colorectal adenocarcinoma cells. This strategy provides an efficient workflow for gene knockout that is rapidly confirmed through PCR amplification of mRNA derived from the targeted gene loci.

Biotechnology and bioengineering↗
PubMed2025

A quick guide to evaluating prime editing efficiency in mammalian cells.

According to the Clinvar database, modeling the diseases associated with pathogenic mutations requires the installation of base substitutions, small insertions or deletions. Prime editor (PE) was recently developed to precisely install any base substitutions and/or small insertions/deletions (indels) in mammalian cells and animals without requiring DSBs or donor DNA templates. PE also offers greater editing and targeting flexibility compared to other precision CRISPR editing methods because the versatile editing information is encoded in the reverse-transcription template of its prime editing guide RNA. However, optimal PE system selection and experimental design can be complex, and there are various factors that can affect PE efficiency. This chapter serves as a rapid entry-level guideline for the application of PE, providing an experimental framework for using PE at a specific genomic locus. RUNX1 was selected as a representative target site to illustrate the detailed methodology for constructing PE plasmids and the process of transfecting these plasmids into 293FT cells. We further examined the efficiency of PE-mediated genome editing in mammalian cells by using next-generation sequencing.

Gene Editing↗
PubMed2026

Base editing reveals an essential role for NANOG in human embryogenesis.

Understanding how the first cell lineages in human development are specified and maintained has fundamental importance and clinical implications for regenerative medicine, infertility and pregnancy loss. Although mouse models have provided valuable insights into transcription factors regulating early development, translating these findings to human embryos has been limited by ethical, technical and biological constraints. Functional studies of transcription factors in human embryos have been hindered by nuclease-based genome editing approaches that induce genotoxicity1-3. Here, to overcome this, we applied ABE8e adenine base editing4,5 to precisely target an exon splice donor site, resulting in a splicing defect and functional knockout of the developmental regulator NANOG in human embryos. This approach did not trigger genotoxicity and showed limited off-target editing. Loss of NANOG disrupts pluripotent epiblast specification and instead cells differentiate towards a primitive endoderm (yolk sac) or trophectoderm (placental) transcriptional programme. Retention of primitive endoderm differentiation in NANOG-edited human embryos reveals a functional compensation that is distinct from mouse, underscoring the importance of directly investigating human development. Our findings demonstrate an essential role for NANOG in human pluripotency and epiblast specification and highlight the utility of base editing for functional interrogation of human development.

Journal Article↗
PubMed2026

Mechanistic Perspectives From Genomics and Pangenomics of Medicinal and Aromatic Plants: Linking Genome Architecture to Phytochemical Diversity.

Medicinal and aromatic plants (MAPs) produce a remarkable diversity of specialized metabolites with significant pharmaceutical, nutraceutical, and industrial value. Although advances in long-read sequencing, chromosome-scale genome assembly, and pangenomics have greatly expanded genomic resources, the mechanistic links between genome architecture and phytochemical diversity remain incompletely understood. The present review synthesizes current evidence describing how structural genomic variation may contribute to phytochemical diversity, while acknowledging that many proposed genome-to-metabolite relationships require further experimental validation. Examples illustrate how genome architecture is associated with specialized-metabolite biosynthesis through multiple regulatory processes. However, the strength of supporting evidence varies considerably among MAP species. Moreover, relatively few genome-to-metabolite relationships have been confirmed through direct functional validation. We further discuss how pangenomics, multiomics integration, genome editing, synthetic biology, and artificial intelligence support the discovery, validation, and engineering of specialized metabolic pathways. Casual conclusions are evaluated according to the strength of available evidence, highlighting where causal relationships have been experimentally established and where conclusions remain primarily association-based. Overall, this review provides an integrated conceptual and evidence-based perspective summarizing proposed relationships between genome architecture and phytochemical diversity and outlines future priorities for functional genomics, precision breeding, metabolic engineering, and sustainable utilization of MAPs.

artificial intelligence↗
PubMed2026

Molecular Bases and Genetic Design of Rice Disease Resistance for Optimized Yield and Sustainable Agriculture.

Rice diseases continue to undermine yield stability and threaten the sustainability of rice production. The central challenge is therefore not simply to maximize immune activation, but to identify genetic interventions that remain effective across diverse pathogen races and environmental conditions without imposing excessive penalties on growth or yield. Here, we synthesize the molecular basis of rice immunity from a design-oriented perspective. We first examine cell-surface pattern-recognition receptors and intracellular nucleotide-binding leucine-rich repeat receptors, and then assess the shared signaling hubs and defence outputs that connect pathogen perception to antimicrobial responses. Rather than treating these components as equivalent breeding targets, we compare their translational potential according to resistance spectrum, anticipated durability, tunability, pleiotropic risk, and the strength of field evidence. We further discuss breeding strategies based on receptor engineering, editing of susceptibility genes and cis-regulatory elements, post-translational motif engineering, pathogen-inducible and upstream open reading frame-mediated regulation, resistance-gene stacking and artificial intelligence-assisted prediction. We argue that rational resistance design in rice should move beyond constitutive immune activation toward allele-specific, quantitative, spatially restricted and infection-responsive regulation. Integrating mechanistic insights with precision genome editing, accelerated breeding and responsible deployment offers a practical route to durable, yield-compatible disease resistance while reducing dependence on chemical control.

breeding strategy↗
PubMed2025

Cell-type-specific response to silicon treatment in soybean leaves revealed by single-nucleus RNA sequencing and targeted gene editing.

Mineral nutrient uptake and deposition profoundly influence plant development, stress resilience, and productivity. Silicon (Si), though classified as a non-essential element, significantly influences a plant's physiology, particularly in fortifying defense responses and mitigating stress. While the genetic and molecular mechanisms of Si uptake and transport are well studied in monocots, particularly rice, their role in dicot species, such as soybean, remains unclear at the cellular and molecular levels. In this study, we utilized single-nucleus RNA sequencing (snRNA-seq) to dissect cellular responses to Si accumulation in soybean leaves. We identified distinct cellular populations, including a unique Si-induced or Si-associated cell cluster within vascular cells, suggesting a specialized mechanism of Si distribution. Si treatment notably induced the expression of defense-related genes, with a pronounced enrichment in vascular cells, underscoring their pivotal role in activating plant defense mechanisms. Moreover, Si modulated the expression of genes involved in phytoalexin biosynthesis, salicylic acid, and immune receptor signaling, suggesting transcriptional priming of genes involved in defense responses. Further investigation of Si transporters revealed precise expression of an Si efflux gene in epidermal cells in response to Si treatment. We also validated the role of efflux Si transporters using a Xenopus oocyte assay and CRISPR/Cas9 genome editing of composite soybean plant roots. This study provides critical insights into the biotic stress regulatory networks influenced by Si treatment in soybean leaves at the single-cell level, thus laying the foundation for enhancing stress tolerance through optimized mineral nutrient uptake.

Glycine max↗
PubMed2026

Miniature and versatile genome regulation TnpB-ωRNA toolkits facilitate cancer immunotherapy.

CRISPR‒Cas systems represent powerful tools for genome regulation. However, the large size of Cas proteins limits their efficient delivery via an adeno-associated virus (AAV), thereby restricting their clinical translation. Here, we engineer the IS200/IS605 transposon-encoded nuclease TnpB, along with its ωRNA scaffold, to create an enhanced TnpB system, which serves as a compact toolkit for gene activation, genome editing, and base editing. The gene activator enTnpBa increases expression by 2889-fold with a minimized 93 nt ωRNA and robustly activates endogenous genes in mammalian cells. We develop a single-AAV-based regimen for immune activation (AAV-ImmunAct) that delivers enTnpBa to activate CXCL9, IL-15, and IFN-γ. AAV-ImmunAct effectively enhances T cell migration and activation, increases killing of cancer cell lines and patient-derived organoids, and synergizes with anti-PD-1 therapy in humanized mice. Here, we establish enTnpB as a compact and versatile platform for genome regulation and a promising tool for cancer immunotherapy.

Humans↗
PubMed2026

Enhanced exonuclease-Cas9 systems promote multiple nucleotide deletions with higher efficiency and broader targeting scope in plants.

CRISPR-Cas9 is a widely used platform for plant genome editing, but its outcomes are typically dominated by small insertions and deletions (indels). Such limited mutation profiles restrict its utility in functional studies of non-coding RNAs and regulatory elements, such as microRNAs (miRNAs), untranslated regions (UTRs), and promoter sequences, where larger sequence disruptions are often required. Here, we developed enhanced exonuclease-Cas9 platforms, termed multiple nucleotide deletion Cas9 (MND-Cas9) systems, for efficient generation of large deletions in rice. By screening four exonucleases (RecJ, T5, TREX2, and SbcB), we established MND-Cas9v1 systems based on TREX2 or SbcB that produced substantially larger deletions without reducing editing efficiency. Further optimization with an inserted DNA-binding domain (DBD) between Cas9 and exonuclease yielded MND-Cas9v2, which simultaneously enhanced efficiency and deletion size. To expand PAM compatibility, we introduced PAM-relaxed Cas9-NG and SpG variants, generating MND-Cas9-NG/SpGv2 systems with broader targeting scope and superior performance compared to their parental nucleases. Finally, we demonstrated the utility of these systems in two applications: MND-Cas9v2 efficiently knocked out the miRNA gene OsMIR530, producing larger seeds, and generated extended deletions in the 3'UTR of OsGhd2, which upregulated its expression and increased grain size. These results demonstrate that MND-Cas9 systems enable high-efficiency generation of extended deletions and facilitate functional analyses of non-coding RNAs and regulatory sequences. Overall, this work establishes a versatile and expandable exonuclease-Cas9 platform that substantially broadens the mutational spectrum and application potential of CRISPR-Cas9 for plant genome engineering.

CRISPR-Cas Systems↗
PubMed2026

Overcoming Immunological Barriers in MSC-Derived Insulin-Producing Cells through CRISPR-Based Hypoimmunogenic Engineering and Translational Perspectives for Type 1 Diabetes.

Mesenchymal stromal cell (MSC)-derived insulin-producing cells (IPCs) represent an emerging strategy for β-cell replacement in type 1 diabetes mellitus (T1DM) owing to their differentiation potential, intrinsic immunomodulatory properties, and lower tumorigenic risk compared with pluripotent stem cell-derived platforms. However, accumulating evidence indicates that differentiation-associated immunogenicity, context-dependent immune recognition, and recurrent autoimmune responses may substantially limit long-term graft survival and therapeutic durability following transplantation. This review critically examines the immunological barriers associated with MSC-derived IPCs, including altered MHC expression, susceptibility to alloimmune and autoimmune-mediated rejection, and potential reactivation of autoreactive immune memory. We discuss the application of CRISPR-based hypoimmunogenic engineering strategies targeting antigen presentation pathways, NK-cell activation, and immune checkpoint modulation to generate more immune-evasive MSC-derived IPCs while preserving β-cell functionality. By integrating insights from T1DM immunopathogenesis, MSC biology, genome editing, and translational immunology, we propose a framework linking immune engineering with controlled differentiation, functional maturation, and long-term safety evaluation. In parallel, we comparatively position MSC-derived IPCs alongside clinically advancing iPSC-derived β-cell platforms to highlight their distinct translational niche, including potential advantages related to safety, immunomodulatory capacity, manufacturing accessibility, and scalability, while acknowledging the superior functional maturity and clinical progression currently demonstrated by iPSC-derived systems. Finally, we discuss key translational challenges, including genomic stability, immune-evasion durability, GMP-compliant manufacturing, and the need for rigorous functional and immunological benchmarking prior to clinical application of hypoimmunogenic MSC-derived IPC therapies in T1DM.

Humans↗
PubMed2026

CRISPGen: A deep generative framework for multi-objective CRISPR/Cas9 guide RNA design via Conditional Latent Diffusion and Dual-Critic Reinforcement Learning.

MOTIVATION: The CRISPR-Cas9 system offers transformative potential for precision genome editing, yet its clinical translation remains constrained by the risk of unintended off-target double-strand breaks. While current discriminative models excel at evaluating pre-specified candidate guides, resolving the fundamental antagonism between on-target cleavage efficiency and off-target specificity within a fixed sequence search space remains a major challenge. RESULTS: We present CRISPGen, a unified deep generative framework that reframes sgRNA design as a multi-objective constrained sequence synthesis problem. It integrates (i) DNABERT-2 genomic-language embeddings, (ii) a conditional latent diffusion generator conditioned on a user-specified on-target efficiency target, and (iii) a dual-critic reinforcement-learning (RL) stage that couples a frozen on-target efficiency critic with a cross-attention off-target discriminator (validation Pearson R=0.8157) trained on a unified corpus of experimental off-target events from six detection platforms. Across 1000 generated sgRNAs, CRISPGen reduces the mean off-target discriminator score by 99.7% relative to the pre-RL baseline and, under an exhaustive whole-genome screen of all 302,631,056 NGG PAM sites in GRCh38, yields zero perfect-match and only 55 one-mismatch genomic hits. We further show, transparently, that the internal on-target critic saturates under RL optimization - an instance of Goodhart's Law - and therefore assess on-target viability using an independent external CRISPRon screen (mean 47.10/100). Repeating the RL fine-tuning stage under three random seeds (with the diffusion generator, DNABERT-2 embeddings, and off-target discriminator held fixed) yields a stable operating point across seeds. Full diversity, per-mismatch, and reproducibility statistics are reported in the Results. AVAILABILITY: Source code is available at https://github.com/malekpouri/CRISPGen; the pre-trained checkpoints and the 3,000,000-sequence library are hosted on Hugging Face (https://huggingface.co/malekpouri/CRISPGen-Checkpoints) and archived on Zenodo under DOI 10.5281/zenodo.21428641.

CRISPR-Cas9↗
PubMed2026

A voyage of reprogrammable metabolic bioengineering reshapes plant defense: from editing tools to synthetic systems.

Metabolic bioengineering has emerged as a transformative approach for reshaping plant defense by targeting intrinsic biosynthetic pathways to enhance immunity in modern agriculture. Moving beyond proof-of-concept metabolomics to broad-spectrum programmable pathway engineering addresses gaps in plant rational design and optimizes resilience in response to diverse environmental cues. This review aims to comprehensively highlight the transition of innovative approaches to phenolics, alkaloids, flavonoids, terpenoids, and benzoxazinoids, inferring adaptive reprogramming that mediates the growth-defense balance and functions as molecular sentinels in plants. Furthermore, decoding the volatile metabolome reveals a dynamic signaling interface that influences defense responses and stress-induced plant-microbe interactions, with the shikimate, jasmonate, and salicylate pathways functioning as central hubs for microbial deterrence and priming immune memory. Recent developments in multi-scalar genome-editing strategies, including CRISPR-driven combinatorial edits, enzyme orthogonalization, fluxomics, and spatially resolved multi-omics, reconfigure central and specialized metabolic fluxes toward improved defense function and regulation. Additionally, emerging tools, such as WUSCHEL2 and BABY BOOM transcriptional modules, and artificial engineering strategies integrating deep learning model-driven predictions facilitate rapid development of synthetic genetic circuits and support a predictive engineering of plants. Moreover, Mass spectrometry imaging (MSI) in spatial metabolomics enables to obtain structures and locations of unidentified endogenous metabolites within cells and tissues. Overall, this review emphasizes a diverse array of primary and secondary metabolites, spanning molecular concepts to recent advances in plant immune mechanisms. It also illustrates new frontiers in programmable metabolic engineering that accelerate the understanding of plant-microbe-metabolite cross-talks, offering strategies to improve plant resistance and advance sustainable agricultural solutions.

metabolic bioengineering↗
PubMed2026

Interfering with DNA repair pathways to enhance CRISPR-Cas9-mediated homology-directed repair in a chelicerate genetic model.

The two-spotted spider mite, Tetranychus urticae, is a major pest and an emerging genetic model. Recent CRISPR-Cas9 advances, especially the SYNCAS method for maternal delivery of Cas9 ribonucleoproteins, have enabled precise genome editing in this and other difficult-to-transform arthropods. Yet SYNCAS-mediated knockins vary in efficiency, possibly due to competition between DNA repair pathways, whose mechanisms in T. urticae and other chelicerates remain unknown. Here, we provide the first functional analysis of double-strand break repair in a chelicerate. Loss of DNA polymerase theta (Polθ) redirects repair almost entirely toward homology-directed repair, whereas absence of Ligase IV has no detectable impact. Using a reporter assay targeting phytoene desaturase, we demonstrate that Polθ-deficient strains enhance incorporation of repair templates, even when mutations are distant from the cut site. Also, insertion of larger fragments is improved. Finally, disrupting Polθ imposes only a modest fitness cost, highlighting its value for future genome engineering in this species.

Acari↗
PubMed2026

Molecular characterization of vitellogenin and its receptor with CRISPR-based sgRNA validation in the legume pod borer, Maruca vitrata (Geyer) (Lepidoptera: Crambidae).

Maruca vitrata, the legume pod borer, causes yield losses of up to 80% in grain legumes. Increasing insecticide resistance and environmental concerns necessitate sustainable pest management alternatives. In the present study, the complete vitellogenin (Vg) coding sequence (CDS), a key reproductive gene involved in oogenesis and embryonic development, was cloned and molecularly characterised from M. vitrata. The assembled Vg CDS (∼5.3 kb) shared 99.04% sequence identity with the reported M. vitrata Vg sequence (MG799570.1). Phylogenetic analysis demonstrated close evolutionary association with related Lepidopteran species, while protein domain analysis identified three conserved domains, namely LPD_N, DUF1943, and VWD. Among these, the single exon-encoded LPD_N domain was selected as the target region for CRISPR/Cas9-mediated editing. Homology models of Vg and vitellogenin receptor (VgR) (Global Model Quality Estimation (GMQE): 0.58 and 0.51) showed a favourable interaction by protein-protein docking (score: -295.66). Three single-guide RNAs (sgRNAs) were designed, synthesised through in-vitro transcription, and evaluated using in vitro cleavage assays. sgRNA1 targeting the LPD_N domain and sgRNA2 targeting the signal peptide region exhibited efficient site-specific cleavage activity, whereas sgRNA3 failed to induce cleavage because of an unfavourable secondary structure that likely impaired Cas9-sgRNA complex formation. Overall, this study provides the first CRISPR-oriented functional characterisation and sgRNA validation of the M. vitrata Vg gene, together with structural characterisation of VgR and Vg-VgR interaction analysis, providing preliminary molecular resources for future CRISPR/Cas9 studies and supporting future embryo microinjection and heritable genome editing for sustainable management of M. vitrata.

CRISPR/Cas9↗
PubMed2026

Reinforcement learning-based dynamic ensemble for missense variant effect prediction and tiered prioritization of VUS.

BACKGROUND: Accurate classification of missense variants remains a challenging task despite major advances in genomics. Numerous computational models have been developed to assist in variant classification, but often require repeated integration and benchmarking efforts. Ensemble methods have been proposed to overcome the limitations of single predictors, but mostly rely on fixed, predefined weights that constrain their ability to capture interactions among predictive signals. METHODS: We present GenixRL, a dynamic ensemble framework that reformulates model fusion as a reinforcement learning optimization problem. GenixRL uses a Q-learning agent to learn a policy that dynamically weights the probabilistic outputs of complementary predictors, including BayesDel (addAF and noAF), ClinPred, and MetaRNN. Replacing static weighting with policy learning allows GenixRL to adaptively identify optimal weightings and substantially improve classification accuracy. RESULTS: In benchmark evaluation against 25 state-of-the-art predictors, GenixRL achieved an AUROC of 0.9644 on an independent ClinVar dataset. On saturation genome editing assays for BRCA1 and BRCA2, GenixRL achieved the best performance and ranked highest on 14 of 17 clinically significant genes in a zero-shot evaluation. Applied to uncertain and conflicting ClinVar variants, GenixRL enabled tiered, evidence-based prioritization of hundreds of thousands of variants as likely pathogenic or pathogenic with high confidence, supported by orthogonal population evidence from gnomAD. CONCLUSION: GenixRL advances pathogenicity prediction for missense variants and provides an adaptive ensemble that sorts variants of uncertain significance into tiered candidates for expert curation and functional validation.

Mutation, Missense↗
PubMed2025

Inactivation of β-1,3-glucan synthase-like 5 confers broad-spectrum resistance to Plasmodiophora brassicae pathotypes in cruciferous plants.

Clubroot disease, caused by the obligate intracellular rhizarian protist Plasmodiophora brassicae, is devastating to cruciferous crops worldwide. Widespread field P. brassicae pathotypes frequently overcome the pathotype-specific resistance of modern varieties, posing a challenge for durable control of this disease. Here a genome-wide association study of 3 years of data comprising field clubroot phenotyping of 244 genome-resequenced Brassica napus accessions identified a strong association of β-1,3-glucan synthase-like 5 (GSL5) with clubroot susceptibility. GSL5 was evolutionarily conserved, and inactivation of GSL5 by genome editing in Arabidopsis, B. napus, Brassica rapa and Brassica oleracea conferred broad-spectrum, high-level resistance to P. brassicae pathotypes without yield penalties in B. napus. GSL5 inactivation derepressed the jasmonic acid-mediated immunity during P. brassicae secondary infection, and this immune repression was possibly reinforced through stabilization of GSL5 by a P. brassicae effector, facilitating clubroot susceptibility. Our study provides durable resistance resources for cruciferous clubroot disease control and insights into plant resistance against intracellular eukaryotic phytopathogens.

Disease Resistance↗