Biology subjects

Zhou, J.

Publications and source records attributed to Zhou, J..

At least 55 records · Page 3Linked to original sources

bioRxiv2026

Decoding Spatial Programs in Human Glioblastoma Through Surprisal Information-Theoretical Analysis

Glioblastoma (GBM) is a highly heterogeneous and invasive brain tumor in which complex interactions among tumor cells, immune cells, and neurons shape disease progression and therapeutic resistance. Resolving spatial patterns programs in glioblastoma (GBM) requires analytical approaches that go beyond variance-driven embeddings. Here, we applied thermodynamic surprisal analysis, an information-theoretic decomposition framework, to spatial transcriptomic sequencing from human GBM specimens to identify orthogonal constraint modes that capture dominant and previously hidden spatial programs. Surprisal analysis revealed structured patterns in the data that are not highlighted by standard approaches such as PCA or cell type deconvolution, including immune-activation-associated signatures as well as spatial programs consistent with synapse remodeling. Coupling surprisal decomposition with spatial ligand-receptor interaction analysis, NICHES, along with multiplexed protein imaging connected these spatial hidden modes to reveal potential communication networks. Together, these results position surprisal analysis as a powerful, complementary strategy for interrogating spatial tumor architecture, enabling discovery of non-obvious spatial programs and interactions that are obscured by variance-based methods.

systems biology↗

bioRxiv2026

Complete elucidation and heterologous reconstruction of the biosynthetic pathway of camptothecin

Camptothecin derivatives are first-line anticancer drugs used worldwide for the treatment of diverse malignant tumors. However, the biosynthetic pathway of camptothecin has remained elusive for five decades. Here, we fully map its entire biosynthetic route. We discovered five key missing enzymes (OpCAR, OpSDR11, OpCS, OpGH1, and OpSTR) via the combination of MALDI mass spectrometry imaging, single-cell RNA sequencing and co-expression analysis. Meanwhile, we demonstrated a free flavin mononucleotide triggered the non-enzymatic 6-5-6 to 6-6-5 fused-ring skeleton rearrangement, filling the last gap in camptothecin biosynthesis. Finally, we validated this identified pathway and achieved the de novo biosynthesis of camptothecin in Saccharomyces cerevisiae. These discoveries uncover the long-standing mystery underlying camptothecin and pave the way for manufacturing camptothecin and its derivatives through synthetic biology approaches.

synthetic biology↗

bioRxiv2026

Microfluidic Platform for Drug Response Profiling in NSCLC Patient-Derived Organoids

Tumor models that recapitulate 3D architecture are essential for understanding how cellular organization and microenvironmental interactions govern therapeutic response in human cancers. Here, we developed a microfluidic microphysiological system that enables controlled and scalable culture and drug testing of non-small cell lung cancer spheroids and patient-derived organoids. The platform integrated U-shaped microwells with dual-channel loading to support de novo spheroid formation, efficient trapping of pre-formed spheroids, and loading of intact organoids with reduced size heterogeneity. Tumor spheroids and organoids maintained high viability and structural integrity during long-term on-chip culture, and constrained microscale confinement produced ellipsoidal geometries that deviate from idealized spherical assumptions. Baseline genotype-dependent responses to KRAS G12C and EGFR inhibitors were preserved across agarose and microfluidic formats, establishing a validated reference state. Building on this baseline, fibroblast- and endothelial-derived cues consistently attenuated responses to targeted therapies across conditioned media, mixed co-culture, and spatially organized configurations. Resistance phenotypes converged on a dominant role for paracrine signaling, while increasing architectural complexity primarily enhanced morphological fidelity rather than altering therapeutic response. These findings establish a microphysiological framework that decouples tumor-intrinsic drug sensitivity from microenvironment-mediated modulation, enabling the systematic evaluation of paracrine resistance mechanisms in NSCLC.

bioengineering↗

bioRxiv2026

A Tripartite Map of the Ventromedial Prefrontal Cortex

The ventromedial prefrontal cortex (VMPFC) has been repeatedly implicated in affect, valuation, and social cognition, yet how these diverse functions are organized within a single cortical territory has remained unresolved. Here, we integrate large-scale meta-analysis, individual-level task fMRI, artificial neural-network encoding models, and multimodal connectivity analyses to reveal the internal functional architecture of the human VMPFC. Across four complementary studies, we identify a robust tripartite organization along the anterior-posterior axis, comprising posterior affective, middle valuation, and anterior social functional motifs. Connectivity fingerprinting demonstrates that each motif is preferentially embedded within distinct large-scale brain networks, providing a mechanistic account of VMPFC functional specialization. This organization is reproducible at the level of individual subjects, generalizes to naturalistic stimuli, extends across development, and shows cross-species correspondence with non-human primates and multiple neurobiological markers. Together, these findings resolve a long-standing organizational question and establish a biologically grounded framework for interpreting VMPFC function.

neuroscience↗

bioRxiv2026

Programming Brain Cell-Type-Selective Delivery In Vivo with Transporter-Guided Therapeutics

Many diseases arise from dysfunction of defined cell populations, yet most therapeutics distribute broadly, limiting efficacy and causing toxicity. We developed ExACT, a platform for cell-type-selective intracellular delivery that exploits membrane transporters. In vivo screening of combinatorial fluorescent small-molecule libraries in mouse brain identified chemistries whose uptake is dictated by endogenous transporter expression, yielding compounds with preferential entry into neurons, astrocytes, pericytes and endothelial cells. One series showed strong selectivity for brain and retinal endothelium, where Slco1a4 mediated uptake. This selectivity principle extended to the human orthologue SLCO1A2, highly expressed in brain endothelium and oligodendrocytes, where it mediated selective uptake in a humanized mouse model and human iPSC-derived oligodendrocytes. Ectopic expression of SLCO1A2 in neurons via gene therapy created a synthetic entry port, conferring ExACT conjugate uptake on otherwise inaccessible cells. Bifunctional compounds linking transporter-targeting motifs to antisense oligonucleotides or small-molecule drugs retained pharmacological activity while conferring transporter-dependent cell-type selectivity, illustrating how transporter diversity can be harnessed for precision pharmacotherapy.

bioengineering↗

bioRxiv2026

GALILEO: Embodied AI scientist for autonomous therapeutic discovery in dynamic membrane systems

Generalizable Agentic Laboratory Intelligence for Learning, Experimentation, and Optimization (GALILEO) closes the prediction-to-intervention gap in therapeutic peptide discovery. Unlike prior AI-scientist systems, GALILEO couples an Observation-Thought-Action-Summary (OTAS) reasoning loop with robotic peptide synthesis and multimodal phenotyping. Candidate peptides are retrieved from a clinically informed peptide prior (CPP) and locally edited through auditable operations. GALILEO autonomously prioritized LRRC8C and SLC25A1 branches and used wet-lab feedback to update target beliefs, sequence policies, assay choices, and mechanism hypotheses. Peptides generated under this framework blocked LRRC8C currents, perturbed osmolyte/redox homeostasis, and promoted tumor-dependent T-cell activation, while SLC25A1 peptides disrupted citrate-export metabolism, reduced extracellular acidosis, and enhanced CD8+ T-cell function. These results establish retrieval-and-editing-based physical learning as a route for auditable, experimentally grounded interventions and transferable membrane-blocker rules.

Molecular Biology↗

bioRxiv2026

STAG2 Maintains Chromatin Compartmentalization and Represses Regulatory Element Contact to Promote Oncogenic Signaling in Muscle Invasive Bladder Cancer

Contrary to other cancer types, stromal antigen 2 (STAG2) expression is associated with shorter survival and an invasive phenotype in muscle invasive bladder cancer (MIBC). As a cohesin complex component, STAG2 regulates genome organization and cell type-specific transcription, yet its mechanistic role in MIBC remains unclear. Here, we uncover mechanisms through which STAG2 coordinates chromatin architecture and gene regulation in MIBC. Modulation of STAG2 rewired chromatin architecture, altering chromatin contacts and compartmentalization, increasing promoter-enhancer interactions, and reducing short-ranged chromatin loops. At specific gene loci, we discovered that STAG2 has context-specific activating and repressive regulatory functions. At the STAG2-activated gene ABCA1, STAG2 maintained A-compartment chromatin and high levels of promoter acetylation, indicators of active transcription. STAG2 KO resulted in loss of promoter acetylation, a shift from A to B compartment chromatin, and increased occupancy of the co-repressor TRIM28, resulting in ABCA1 downregulation and diminished invasive potential. Conversely, at the STAG2-repressed gene SPOCK3, STAG2 KO resulted in B to A compartment switching, aberrant formation of chromatin loops, and SPOCK3 upregulation. Treatment with EZH2 inhibitor tazemetostat augmented STAG2-KO induced SPOCK3 upregulation, suggesting a collaborative role of STAG2 and EZH2 in repressing Polycomb Repressive Complex 2 (PRC2) target genes. Altogether, our results indicate that STAG2 plays a multifaceted role in regulating gene expression in bladder cancer that is dictated by the epigenetic and chromatin landscape of the cells. These findings identify STAG2-dependent vulnerabilities and provide a rationale for therapeutic targeting of chromatin regulators in MIBC.

Cancer Biology↗

bioRxiv2026

Metal Coordination Dynamics Governs Selective Epoxidation in Hyoscyamine 6β-Hydroxylase: Integrated Experimental and Computational Insights

Iron(II)- and 2-oxoglutarate-dependent (Fe(II)/2OG) enzymes catalyze a wide range of C-H bond activation and functionalization reactions and play essential roles in biosynthesis and metabolic regulation. Despite extensive mechanistic studies, the principles governing selectivity between canonical hydroxylation and alternative transformations remain incompletely understood. Here, we investigate the catalytic mechanism of hyoscyamine 6{beta}-hydroxylase (H6H), a Fe(II)/2OG-dependent oxygenase that sequentially catalyzes the 6{beta}-hydroxylation of hyoscyamine followed by 6,7-exo-epoxidation of 6{beta}-hydroxyhyoscyamine to generate scopolamine. Combined molecular dynamics and QM/MM calculations reveal that an inline Fe(IV)-oxo intermediate initiates hydrogen atom abstraction from the substrate C7 position. The resulting Fe(III)-OH species subsequently deprotonates the substrate hydroxyl group in a process coupled to substrate coordination to the iron center and an in-line-to-off-line rearrangement of the Fe(III)-OH moiety. This coordination dynamics machinery is further supported by the observed chlorination reactivity on the same substrate. Importantly, this coordination switch favors epoxide formation over hydroxyl rebound, thereby directing the reaction toward selective epoxidation. Further computational analysis of the L290F variant demonstrates that steric constraints imposed by L290 are essential for suppressing hydroxylation, revealing a bidirectional regulatory mechanism governing epoxidation/hydroxylation selectivity. Whereas iron coordination dynamics promote epoxidation reactivity, precise substrate positioning and protein-derived steric effects suppress the competing hydroxylation pathway. These findings are consistent with available experimental observations and establish metal coordination dynamics as a key determinant of selective C-H functionalization in Fe(II)/2OG enzymes.

biochemistry↗

bioRxiv2026

Prenatal assembly of functional cortical circuits

The extent to which early brain maturation requires interactions with the outside world is a central question in neurobiology. Because much of cortical maturation occurs after birth in Mus, it has often been viewed as dependent on postnatal experience. However, studies addressing this issue have largely relied on postnatal sensory deprivation paradigms, which perturb normal development and cannot determine to what extent postnatal experience per se drives maturation. To address this question, here we compare two related rodents with markedly different gestation lengths: the precocial Acomys dimidiatus (39-day gestation) and the altricial Mus musculus (19-day gestation). By generating a novel Acomys reference genome and using histology, cellular birth dating, electrophysiology, single-nucleus transcriptomics, and quantitative behavior, we show that Acomys preserves the canonical sequence and timing of cortical development, while shifting major milestones of neuronal, circuit and behavioral maturation into prenatal life. At birth, Acomys cortex already shows advanced cytoarchitecture, neuronal physiology, thalamocortical barrels, transcriptional states, and sensorimotor behavior, with postnatal molecular programs in Mus unfolding prenatally in Acomys. Thus, birth is not a prerequisite for early cortical maturation. Instead, evolutionarily conserved developmental programs unfold across birth, with birth occurring at different stages of these programs, reflecting a species-specific balance between neonatal competence and prolonged postnatal plasticity.

neuroscience↗

bioRxiv2026

Intron 13 retention expands the ADAR1 isoform repertoire to rewire PKR signaling and promote tumorigenesis

For decades, the biology of ADAR1 has been framed around two major isoforms: the nuclear-enriched p110 and the predominantly cytoplasmic p150. Here, we reveal an unexpected repertoire expansion of ADAR1 isoforms that is driven by intron retention. Specifically, intron 13 (I13) can be retained in ADAR1 transcripts, conferred by an evolutionarily conserved weaker 5SS. In addition, we found that the I13 retention (I13R) is negatively autoregulated by ADAR1 through antagonizing the binding of hnRNPA1 to I13 in an editing-independent manner. Despite being sensitive to nonsense-mediated decay, I13R generates two previously uncharacterized truncated isoforms - p90 and p130, that both lack the C-terminal portion of the deaminase domain. Intriguingly, ADAR1p90 - a derivative of the canonical nuclear-enriched p110 isoform - is predominantly cytoplasmic, that effectively represses PKR and eIF2 activation through the sequestration of immunogenic double-stranded RNA (dsRNA) substrates. Furthermore, in a colorectal cancer (CRC) cohort, p90 levels are increased in most tumors relative to matched normal tissues that positively correlates with hnRNPA1 expression. Functionally, xenografts expressing ADAR1p90 grow significantly faster and larger than those expressing ADAR1p110, indicating enhanced tumorigenic potential. These findings revise the canonical view of ADAR1 isoforms, demonstrating that intron retention can generate alternative isoforms with augmented functions that tumors readily exploit. One Sentence SummaryThe conserved retention of I13 in ADAR1 transcripts gives rise to previously uncharacterized C-terminal truncated cytoplasmic ADAR1 isoforms, p90 and p130, which, although devoid of catalytic activity, sequester immunogenic dsRNA substrates, thereby preventing PKR binding and downstream activation of eIF2.

cell biology↗

bioRxiv2026

Revisiting CPUs for Protein Folding: Xeon-Based Acceleration of AlphaFold2

Protein structure prediction via AlphaFold2 has revolutionized drug discovery, yet its end-to-end execution remains computationally intensive. While GPUs are traditionally favored for deep learning, the AlphaFold2 algorithm consists of heterogeneous phases -- preprocessing with sparse database searches and model inference with low-arithmetic-intensity attention modules -- that present unique architectural challenges. In this work, we address these bottlenecks by introducing Open-Omics-AlphaFold2, a highly optimized implementation for Intel(R) Xeon(R) CPU. By leveraging the CPUs versatility in handling both sparse preprocessing algorithms and dense matrix operations via Intel Advanced Matrix Extensions (AMX), we accelerate the entire pipeline end-to-end. Our optimization strategy employs multi-level parallelism -- spanning multiprocessing, multi-threading, and vectorization -- alongside cacheaware tiling and operator fusion. Our results demonstrate that, on a Xeon CPU, Open-Omics-AlphaFold2 achieves 2 7.58 speedup for preprocessing and 19.8 29.2 speedup for model inference over baseline Deepmind-AlphaFold2. Moreover, for a proteome of 391 proteins, Open-Omics-AlphaFold2 running on a dual-socket Intel Xeon 6980P system achieves a remarkable 76% higher through-put over the state-of-the-art GPU-accelerated solution, FastFold, running on a single-socket Intel Xeon 6980P CPU with an NVIDIA H100 offioad. Code availabilityBaremetal: https://github.com/IntelLabs/open-omics-alphafold Containerized: https://github.com/IntelLabs/Open-Omics-Accelera tion-Framework/tree/main/pipelines/alphafold2-based-protein-folding

genomics↗

bioRxiv2026

Fungal community composition links rhizosphere microbiome organization to plant phenotype in response to moderate warming

BackgroundGlobal warming increasingly challenges plant performance and ecosystem function, and plant responses to elevated temperature are shaped not only by intrinsic plasticity, but also by interactions with rhizosphere microbial communities. However, it remains unclear how warming-induced microbiome reorganization relates to plant performance, whether microbiome community composition predicts plant phenotypic responses to elevated temperature better than simple host-microbiome origin matching, and which bacterial or fungal community features contribute most strongly to this prediction. ResultsWe implemented a full factorial design manipulating plant genotype (P), microbial inoculum (M), and temperature (T) using natural Arabidopsis thaliana ecotypes and their corresponding rhizosphere microbiomes from contrasting climatic regions. By integrating high-throughput plant phenotyping, microbial community profiling, microbiome-phenotype coupling analyses, and predictive modeling, we found that elevated temperature induced a coherent thermomorphogenic shift in plant architecture, while plant phenotypic variation remained determined primarily by genotype over the 14-day vegetative growth period covered in this study. Overall, cold-origin genotypes showed larger temperature-induced trait shifts than warm-origin genotypes. Rhizosphere microbial communities were structured predominantly by inoculum origin, but were also affected by warming. Evidence for a increased thermomorphogenic growth when genotypes were combined with inoculum from their home site was limited. Instead, microbiome-phenotype associations were better captured by community composition than by matching status, with fungal community variation showing stronger and more consistent associations with plant phenotype than bacterial variation and providing more robust predictions of plant phenotypes across models. ConclusionsWarming reorganized plant-rhizosphere systems within persistent host- and inoculum-associated baselines. Plant phenotypic variation during vegetative development under elevated temperature was linked more closely to microbiome composition, especially in fungi, than to simple host-microbiome matching of origins. These findings provide a framework for identifying microbiome features associated with plant performance under climate warming.

plant biology↗

bioRxiv2026

Rapid in vitro synthesis of DNA templates via Sidewinder for polyadenylated Hantavirus mRNA vaccine candidates

As the recent COVID-19 pandemic illustrated, zoonotic viruses and other pathogens pose a credible threat to public health. Recent advancements in vaccine technology, particularly mRNA vaccines, provide key tools for an effective and swift public health response. Although mRNA vaccines can be developed more quickly than traditional vaccines, fast and accurate construction of DNA templates for these vaccines remains a critical bottleneck. Using our novel DNA assembly technology, Sidewinder, we rapidly designed and built multiple mRNA vaccine candidates to guard against a potential outbreak of Hantavirus (ANDV). We successfully constructed the DNA templates from oligo pools and produced the mRNA for three vaccine candidates in just 2 days after delivery of the synthetic DNA oligos.

biochemistry↗

bioRxiv2026

A harmonized single-cell RNA-seq atlas of human localized and metastatic prostate cancers and benign tissues

Single-cell RNA sequencing (scRNA-seq) effectively captures the differences in transcriptomic landscape of cell types and cell states between benign and cancer tissues. Pooling publicly available datasets distributed across independent studies enables increased sample representation and cross-study comparisons. Here we present a harmonized scRNA-seq atlas of the human prostate constructed by integrating 17 available studies, comprising 163 samples from 106 donors. The dataset contains benign tissue, primary tumors, and metastatic disease profiles. Raw sequencing FASTQ data files were uniformly reprocessed to minimize technical variability. Study metadata were curated and standardized using a unified schema capturing donor identity, tissue site, disease context, and histologic grade. Post quality control, the integrated dataset contains 754,000 high-quality cells. Harmonized cell type annotations were generated using a pseudobulk correlation framework informed by multiple reference resources. The workflow identified 17 distinct cell types representing epithelial, mesenchymal, and immune compartments of the prostate. The processed expression matrices, standardized metadata, and analysis workflows are publicly available to support reproducible analysis and enable exploration of heterogeneity across prostate disease states.

cancer biology↗

bioRxiv2026

Humanization of N-glycan-dependent protein quality control system in Kluyveromyces marxianus promotes glycoprotein secretion

Human N-glycoproteins constitute a market worth hundreds of billions of dollars. However, their production in yeast is often limited by misfolding and subsequent degradation, largely due to differences in N-glycan-dependent protein quality control (QC) systems between humans and yeast. Notably, yeast lacks the UGGT-mediated reglucosylation-refolding cycle that rescues misfolded glycoproteins, and its degradation pathway involves fewer rate-limiting steps. To address this, we engineer the glycoprotein QC system in Kluyveromyces marxianus, a promising host for protein production, by introducing key human components and modifying native pathways. Expression of human UGGT1 or UGGT2 enhances the soluble and secretory production of glycoproteins in an activity-dependent manner. This effect is further improved by co-expression of the UGGT cochaperone SEP15 and by reducing native glucosidase II trimming activity. In addition, introduction of human EDEM2, a rate-limiting enzyme in glycoprotein degradation, delays ER-associated degradation and increases secretion. Integration of these engineering strategies substantially enhances the production of several high-value human-derived glycoprotein therapeutics, including etanercept, dulaglutide, and abatacept, with up to a [~]12-fold increase. These findings demonstrate that engineering a human-like glycoprotein QC network in yeast is an effective strategy to improve glycoprotein folding and secretion.

bioengineering↗

bioRxiv2026

Targeting cancer-associated cell surface RNAs with oligonucleotide-drug conjugates enables broad antitumor activity

Recent studies have revealed the presence of RNAs on mammalian cell surfaces, yet the linkage of cell surface RNAs (csRNAs) to cellular states and their potential as extracellular accessible drug targets remains unexplored. Here we develop Cell-surface and Intracellular RNAs Co-mapping (CIRCmap), a highly multiplexed in situ profiling approach that simultaneously detects thousands of cell-surface and intracellular RNAs at single-cell resolution. Using CIRCmap, we uncover that csRNA distributions are correlated with cellular states and identified cancer-associated csRNAs. Integrative analysis of cell-surface and intracellular RNAs within the same cells implies that the csRNAs undergo endocytosis and endolysosomal trafficking, which is further supported by perturbation experiments and co-localization visualization. We design oligonucleotide-drug conjugates (ODCs) that target cancer-associated csRNAs for endocytic delivery of cytotoxic payloads selectively to cancer cells. ODCs exhibit broad antitumor activity in cell lines and an in vivo mouse tumor model, opening a promising new avenue for targeted cancer therapy.

cell biology↗

bioRxiv2026

A mixer tap mechanism in Mycobacterium tuberculosis regulates a critical metabolic node in response to nutrient availability

Mycobacterium tuberculosis persists within the host by exploiting metabolic flexibility underpinned by the tricarboxylic acid (TCA) cycle-glyoxylate shunt junction, where isocitrate dehydrogenase (ICD) and isocitrate lyase (ICL) compete to direct carbon flux between energy production and carbon conservation. Yet, how nutrient availability regulates flux partitioning at this junction is unknown. We show that among the four gatekeeping isoforms of isocitrate lyase (ICL1/2) and isocitrate dehydrogenase (ICD1/2), only icl1 transcription substantially responds to carbon substrate switching. Systematic metabolite screening revealed that pyruvate, oxaloacetate, and glyoxylate are novel allosteric activators of ICD2, while concurrently inhibiting ICL1/2 at their physiological concentrations. To resolve how these regulatory layers integrate, we developed an experimental 1H NMR-based flux assay that combines recombinant enzymes with metabolomics-informed metabolite conditions to quantify flux partitioning under defined states. This systems-level framework reveals that flux partitioning emerges from coordinated balancing of enzyme abundance and protein-metabolite effector combinations that shift in a time-dependent, nutrient-responsive manner. This dynamic, multi-layered regulatory framework, analogous to a mixer-tap mechanism, offers new strategies for therapeutic disruption of M. tuberculosis metabolism.

biochemistry↗

bioRxiv2026

Bipartite DNA binding domain of transcription factor BCL11B binds clustered short DNA sequence motifs

B-cell leukemia/lymphoma 11B (BCL11B), despite its name, is a key regulator of T-cell development, specification, and T-cell malignancies. BCL11B contains a bipartite DNA binding domain composed of two C2H2 zinc finger arrays: low-affinity ZF2-3 and high affinity ZF4-6. These arrays function as homotypic modules that recognize similar six-nucleotide motifs, TG(O_SCPLOWNC_SCPLOW)CC(O_SCPLOWCC_SCPLOWO_SCPCAP/C_SCPCAPO_SCPLOWTC_SCPLOWO_SCPCAP/C_SCPCAPO_SCPLOWAC_SCPLOW), as seven of the eight DNA base-contacting residues are conserved between them. The most conserved interactions involve GG dinucleotides, contacted by arginine and lysine residues at key base-interacting positions in ZF3 and ZF5. The two ZF arrays are connected by a long [~]300-residue linker that provides flexibility in how the arrays engage DNA, allowing ZF2-3 and ZF4-6 binding to the same or opposite strands with variable orientation, spacing and positioning along the DNA. This extended linker is enriched in serine/threonine, acidic residues (aspartate/glutamate), and structural residues (glycine/proline), providing additional layers of transcriptional regulation possibly through post-translational modification, electrostatic modulation, and/or condensate formation. We also examined six missense mutations in base-interacting residues, that are associated with neurodevelopmental disorders. Substitutions replacing bulky, positively charged arginine or lysine with smaller or hydrophobic residues likely reduce DNA-binding affinity and/or specificity, whereas substitutions between asparagine and lysine may alter base recognition preferences.

biochemistry↗