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PubMed2025

An allelic resolution gene atlas for tetraploid potato provides insights into tuberization and stress resilience.

Tubers are modified underground stems that enable asexual, clonal reproduction and serve as a mechanism for overwintering and avoidance of herbivory. Tubers are wide-spread across angiosperms with some species such as Solanum tuberosum L. (potato) serving as a vital crop for human consumption. Genes responsible for tuber initiation and disease resistance have been characterized in potato including StSP6A, a homolog of Flowering Time, that functions as tuberigen, the equivalent of florigen. To elucidate additional molecular and genetic mechanisms underlying potato biology including tuber initiation, tuber development, and stress responses, we generated a developmental and abiotic/biotic-stress gene expression atlas from 34 tissues and treatments of Atlantic, a tetraploid cultivar. Using the haplotype-phased tetraploid Atlantic genome assembly and expression abundances of 129,218 genes, we constructed gene coexpression modules that represent networks associated with distinct developmental stages as well as stress responses. Functional annotations were given to modules and used to identify genes involved in tuberization and stress resilience. Structural variation from a pan-genomic analysis across four cultivated potato genome assemblies as well as domestication and wild introgression data allowed for deeper insights into the modules to identify key genes involved in tuberization and stress responses. This study underscores the importance of transcriptional regulation in tuberization and provides a comprehensive framework for future research on potato development and improvement.

Journal Article↗
PubMed2025

Genome-wide association meta-regression identifies stem cell lineage orchestration as a key driver of acne risk.

Over 85% of the population experience acne at some point in their lives, with its severity spanning a quantitative spectrum, from mild, transient outbreaks to more persistent, severe forms of the condition. Moderate to severe disease poses a substantial global burden arising from both the physical and psychological impacts of this highly visible condition. The analytical approach taken in this study aimed to address the impact of variation in the dichotomisation of acne case control status, driven by ascertainment and study design, on effect size estimates across independent genetic association studies of acne. Through a fixed intercept meta-regression framework, we combined evidence genome-wide for association with acne across studies in which case-control status had been ascertained in different settings, allowing for different severity threshold definitions. Across a combined sample of 73,997 cases and 1,103,940 controls of European, South Asian and African American ancestry we identify genetic variation at 165 genomic loci that influence acne risk. There is evidence for both shared and ancestry specific components to the genetic susceptibility to acne and for sex differences in the magnitude of effect of risk alleles at three loci. We observe that common genetic variation explains 13.4% of acne heritability on the liability scale. Consistent with the hypothesis that genetic risk primarily operates at the level of individual pilosebaceous units, a polygenic score derived from this case-control study of acne susceptibility is associated with both self-reported and clinically assessed acne severity in adolescence, further strengthening the link between genetic risk and disease severity. Prioritisation of causal genes at the identified acne risk loci, provides genetic validation of the targets of established and emerging acne therapies, including retinoid treatments. The identified acne risk loci are enriched for genes encoding downstream effectors of RXRA signalling, including SOX9 and components of the WNT and p53 pathways. Illustrating that the control of stem cell lineage plasticity and cellular fate are important mechanisms through which genetic variation influences acne susceptibility within the pilosebaceous unit.

Journal Article↗
PubMed2025

MarkerMatch: A Proximity-Based Probe-Matching Algorithm for Joint Analysis of Copy-Number Variants from Different Genotyping Arrays.

MOTIVATION: Copy-number variants (CNVs) are a form of genetic structural variation with increasing importance in complex human disorders. Both DNA sequencing and microarray data can be used to call CNVs, which can be used in association tests, such as association between CNV number and disease status. Unlike genotypes, CNV detection in microarrays requires the use of observed intensity signals at each probe, which limits the imputability for analyses that span multiple array types. Thus far, a consensus set of probes (the intersection encompassing the probes that occur in common on all arrays) has been used to circumvent the problem of differing array-specific sensitivities. This has, however, led to excessive reduction in overall sensitivity of CNV calls as arrays can have an undesirably low overlap of probe sets. To overcome this limitation, we developed MarkerMatch, a proximity-based algorithm that matches probes across different genotyping microarrays to maximize the number of probes considered in the CNV calling algorithm, thereby increasing the resolution and sensitivity while preserving precision. RESULTS: By analyzing CNV calls from 4,906 individuals genotyped across three different arrays (Global Screening Array, Omni2.5 array, and Omni Express Exome array), we show that the MarkerMatch approach improves sensitivity by increasing the density of probes available for CNV calling while maintaining precision or improving it relative to the current practice (e.g., use of consensus probes only). We further demonstrate that MarkerMatch exceeds the output from current practice in terms of F1 score, Fowlkes-Mallows index, and Jaccard index. We also optimize MarkerMatch parameters, D MAX and Method, and find an optimal D MAX setting at 10kb, with no clear optimal candidate based on Method, indicating that parameters for this metric should be determined on a use case basis.

Journal Article↗
PubMed2025

Polyamines buffer labile iron to suppress ferroptosis.

Polyamines are essential and evolutionarily conserved metabolites present at millimolar concentrations in mammalian cells. Cells tightly regulate polyamine homeostasis through complex feedback mechanisms, yet the precise role necessitating this regulation remains unclear. Here, we show that polyamines function as endogenous buffers of redox-active iron, providing a molecular link between polyamine metabolism and ferroptosis. Using genome-wide CRISPR screens, we identified a synthetic lethal dependency between polyamine depletion and the key ferroptosis suppressor, GPX4. Mechanistically, we show that polyamine deficiency triggers a redistribution of cellular iron, increasing the labile iron pool and upregulating ferritin. To directly visualize this iron buffering in living cells, we developed a genetically encoded fluorescent reporter for redox-active iron. Live-cell analysis revealed a striking inverse correlation between intracellular polyamine levels and redox-active iron at single-cell resolution. These findings reposition polyamines as key regulators of iron homeostasis, with implications for ferroptosis-linked disease states and cellular redox balance.

Journal Article↗
PubMed2025

The mouse gut microbiota responds to predator odor and predicts host behavior.

Chronic stressors can alter the mammalian gut microbiota in ways that mediate host stress responses, but the impacts of acute stressors on these interactions are less well understood. Here, we show that brief exposure of wild-derived mice to predator odor altered gut-microbiota composition, which in turn predicted host behavior. We investigated the individual and combined effects of 15-minute exposures to synthetic fox fecal odor and 30 days of chronic social isolation, an established chronic stressor. Using ethological assays, visceral adipose tissue transcriptomics, and genome-resolved metagenomics, we found that predator-odor exposure significantly affected mouse behavior, gene expression, and gut microbiota. Predator odor-responsive bacteria were associated with the expression of genes involved in anti-microbial defense, and host behavioral responses were predicted by random forest models trained on gut-microbiota profiles. These findings indicate interactions between the gut microbiota and wild-mouse responses to the threat of predation, an ecologically relevant acute stressor.

Journal Article↗
PubMed2025

Perplexity as a Metric for Isoform Diversity in the Human Transcriptome.

Long-read sequencing (LRS) has revealed a far greater diversity of RNA isoforms than earlier technologies, increasing the critical need to determine which, and how many, isoforms per gene are biologically meaningful. To define the space of relevant isoforms from LRS, many existing analysis pipelines rely on arbitrary expression cutoffs, but a single threshold cannot accommodate the broad variability in isoform complexity across genes, cell-types, and disease states captured by LRS. To address this, we propose using perplexity-an interpretable measure derived from entropy-that quantifies the effective number of isoforms per gene based on the full, unfiltered isoform ratio distribution. Calculating perplexity for 124 ENCODE4 PacBio LRS datasets spanning 55 human cell types, we show that it provides intuitive assessments of isoform diversity and captures uncertainty across genes with varying complexity. Perplexity can be calculated at multiple gene regulatory levels-from transcript to protein-to compare how isoform diversity is reduced across stages of gene expression. On average, genes have an ORF-level perplexity of 2.1, indicating production of two distinct protein isoforms. We extended this analysis to evaluate expression variation across tissues and identified 4,593 ORFs across 3,102 genes with moderate to extreme tissue-specificity. We propose perplexity as a consistent, quantitative metric for interpreting isoform diversity across genes, cell types, and disease states. All results are compiled into a community resource to enable cross-study comparisons of novel isoforms.

Journal Article↗
PubMed2025

Gene model for the ortholog of Glys in Drosophila ananassae.

Gene model for the ortholog of Glycogen synthase (Glys) in the May 2011 (Agencourt dana_caf1/DanaCAF1) Genome Assembly (GenBank Accession: GCA_000005115.1) of Drosophila ananassae. This ortholog was characterized as part of a developing dataset to study the evolution of the Insulin/insulin-like growth factor signaling pathway (IIS) across the genus Drosophila using the Genomics Education Partnership gene annotation protocol for Course-based Undergraduate Research Experiences.

Bioinformatics↗
PubMed2025

mamp-ml: A deep learning approach to epitope immunogenicity in plants.

Eukaryotes detect biomolecules through surface-localized receptors, key signaling components. A subset of receptors survey for pathogens, induce immunity, and restrict pathogen growth. Comparative genomics of both hosts and pathogens has unveiled vast sequence variation in receptors and potential ligands, creating an experimental bottleneck. We have developed mamp-ml, a machine learning framework for predicting plant receptor-ligand interactions. We leveraged existing functional data from over two decades of foundational research, together with the large protein language model ESM-2, to build a pipeline and model that predicts immunogenic outcomes using a combination of receptor-ligand features. Our model achieves 73% prediction accuracy on a held-out test set, even when an experimental structure is lacking. Our approach enables high-throughput screening of LRR receptor-ligand combinations and provides a computational framework for engineering plant immune systems.

Journal Article↗
PubMed2025

Differential expression of a disease-associated MRE11 variant reveals distinct phenotypic outcomes.

The MRE11 DNA nuclease plays central roles in the repair of DNA double-strand breaks (DSBs) as a core component of the heterotrimeric MRE11/RAD50/NBS1 (MRN) complex. MRN localizes to chromosomal DSBs and recruits and activates the apical DSB repair protein kinase, ATM, which phosphorylates downstream substrates to elicit cellular DNA damage responses. Pathogenic variants in MRE11 cause the genome instability disorder ataxia-telangiectasia-like disorder (ATLD). The first ATLD patient allele identified, ATLD1, is a nonsense mutation that deletes 76 amino acids from the MRE11 C-terminus and results in markedly reduced levels of MRE11-ATLD1 and the entire MRN complex. This region of the C-terminus has been demonstrated to function in DNA binding, mediate functional protein interactions, and undergo post-translational modifications that regulate MRE11 nucleolytic activities. We previously demonstrated that transgenic mice expressing low wildtype MRN exhibit severe phenotypes, including small body size, anemia, and cellular DNA DSB repair defects. Thus, it is currently unknown whether reduced MRE11-ATLD1 and MRN levels, loss of the C-terminus, or both cause disease-associated phenotypes. In this study, we generated transgenic mouse models that express near endogenous or significantly reduced levels of MRE11-ATLD1 to determine the in vivo importance of the MRE11 C-terminus. We observe that reduced MRE11-ATLD1 expression leads to anemia, bone marrow failure, extramedullary hematopoiesis, and impaired lymphocyte development, similar to mice expressing low wildtype MRE11. In contrast, higher expression of MRE11-ATLD1 results in a subset of moderate phenotypes, indicating that loss of C-terminus has limited impact on MRN functions in vivo. These findings have implications for clinical predictions of ATLD patients harboring pathogenic MRE11 variants that impair MRE11 function and/or impact MRN protein levels.

Journal Article↗
PubMed2025

Pervasive noise in human splice site selection.

RNA splicing has historically been thought to be highly efficient and accurate, with little opportunity for deviation from regulated alternative splicing decisions. This dogma has been challenged by recent observations that suggest that biological noise may contribute substantially to transcriptome diversity. However, quantitative understanding of stochastic variations in splicing is challenging because these transcripts are likely subject to rapid degradation. Here, we use ultra-deep sequencing across RNA compartments to track splicing intermediates in human cells and see abundant cryptic splicing associated with genomic features that promote splicing noise. We observe pervasive usage of low-fidelity splice sites, likely due to stochasticity in recruitment or binding of the spliceosome. These sites are most likely degraded in the nucleus rather than targeted by translation-dependent degradation processes, suggesting widespread surveillance and rapid quality control of non-productive RNA transcripts. Our findings provide unprecedented insights into the propensity for error in RNA processing mechanisms and the regulation of alternative splice sites across a gene.

Journal Article↗
PubMed2025

Gene model for the ortholog of Pi3K21B in Drosophila eugracilis.

Gene model for the ortholog of Phosphatidylinositol 3-kinase 21B (Pi3K21B) in the D. eugracilis May 2021 (Stanford ASM1815383v1/DeugRefSeq2) Genome Assembly (GenBank Accession: GCF_018153835.1) of Drosophila eugracilis. This ortholog was characterized as part of a developing dataset to study the evolution of the Insulin/insulin-like growth factor signaling pathway (IIS) across the genus Drosophila using the Genomics Education Partnership gene annotation protocol for Course-based Undergraduate Research Experiences.

Journal Article↗
PubMed2026

Intersecting experimental evolution and CRISPR screens to identify novel toxin resistance loci.

Understanding toxin resistance in insects is key to appreciate niche adaptations but remains challenging due to its often-polygenic basis. A well-known example is the specialized association of Drosophila sechellia with noni fruit ( Morinda citrifolia ), which is toxic to most other insects, including the closely-related Drosophila simulans and Drosophila melanogaster . Toxicity of noni is due to its high concentration of octanoic acid (OA), but the mechanisms that determine sensitivity or resistance to OA in different species remain poorly understood. Here, we experimentally-evolved D. simulans with increased OA resistance, identifying multiple loci under selection. Cross-referencing these with a genome-wide, OA-resistance CRISPR screen in a D. melanogaster cell line highlighted two proteins: Kraken, a putative detoxification enzyme expressed in digestive and renal tissues, and Alkbh7, a mitochondrial protein linked to fatty acid metabolism. Both genes show elevated expression in D. sechellia and OA-resistant D. simulans . In D. melanogaster , kraken mutants are more OA-sensitive, while Alkbh7 overexpression increased OA resistance. Importantly, mutation of these genes in D. sechellia reduced OA tolerance. Our identification of genes underlying OA resistance in laboratory and natural contexts demonstrates how complementary, cross-species selection approaches can provide insights into complex mechanisms of toxin susceptibility and adaptation; such methods could also have practical applications in the characterization of natural and artificial insecticides.

Journal Article↗
PubMed2025

Differential Crosslinking and Contractile Motors Drive Nuclear Chromatin Compaction.

During interphase, a typical cell nucleus features spatial compartmentalization of transcriptionally active euchromatin and repressed heterochromatin domains. In conventional nuclear organization, euchromatin predominantly occupies the nuclear interior, while heterochromatin, which is approximately 50% more dense than euchromatin, is positioned near the nuclear periphery. Peripheral chromatin organization can be further modulated by the nuclear lamina, which is itself a deformable structure. While a number of biophysical mechanisms for compartmentalization within rigid nuclei have been explored, we study a chromatin model consisting of an active, crosslinked polymer tethered to a deformable, polymeric lamina shell. Contractile motors, the deformability of the shell, and the spatial distribution of crosslinks all play pivotal roles in this compartmentalization. We find that a radial crosslink density distribution, even with a small linear differential of higher crosslinking density at the edge of the nucleus, combined with contractile motor activity, drives genomic segregation, in agreement with experimental observations. This arises from contractile motors preferentially drawing crosslinks into their vicinity at the nuclear periphery, forming high-density domains that promote heterochromatin formation. We also find an increased stiffness of nuclear wrinkles given the preferential heterochromatin compaction below the lamina shell, which is consistent with instantaneous nuclear stiffening under applied nanoindentation. We conclude with the potential for experimental validation of our model predictions.

Journal Article↗
PubMed2025

A tumorigenesis threshold for endogenous Myc revealed by dosage-compensation for Myc-haploinsufficiency in the absence of p53.

The MYC proto-oncogene is crucial for neoplasia in most tumors. Overexpressed, oncogenic MYC amplifies the flux through most major processes but does not specify a unique carcinogenic pathway. This "amplifier" model suggests that MYC must exceed an expression threshold to become oncogenic. We designed a genetic test of this model, using the mouse Trp53 null mutant (p53KO) as a highly robust tumor generator to examine the effect of a modest change in the endogenous Myc level (Myc +/-). Strikingly, tumor-free survival is greatly extended in p53KO mice with haploid Myc gene-dosage, yet in the tumors that do develop (mainly hemangiosarcomas and thymic lymphomas), their Myc deficit has been invariably compensated either by increasing Myc genomic dosage (hemangiosarcomas) or expression (lymphomas). Furthermore, acutely halving the endogenous Myc gene-dosage in established tumor allografts curtails growth rates. These results indicate that even an incremental reduction of MYC activity can be salutary in cancer and that one of the major tumor suppressor functions of p53 derives from its ability to prevent MYC overexpression. Myc generates acute DNA damage by several mechanisms and accordingly, p53's anti-Myc function may be inextricably linked to its role in genome integrity surveillance.

Myc haplo-insufficiency↗
PubMed2025

A xeno-free protocol for rapid differentiation of human iPSC-derived microglia from the KOLF2.1J reference line.

We present a detailed, xeno-free protocol for the rapid differentiation of human induced pluripotent stem cells (hiPSCs) into microglia using the well-characterized KOLF2.1J reference line. This system employs doxycycline-inducible expression of six transcription factors (6-TF), stably integrated into the CLYBL safe harbor locus, to drive uniform microglial differentiation within two weeks. Adapted from Dräger et al. (1), our protocol includes key optimizations for KOLF2.1J, including culture on Laminin-521 to support xeno-free conditions. The resulting i-Microglia exhibit hallmark features of mature microglia, including expression of P2RY12, loss of the pluripotency marker SSEA4, phagocytic activity, and upregulation of immune markers (e.g., CD80, CD83) upon LPS stimulation. We also demonstrate compatibility with co-culture systems using iPSC-derived neurons. Additionally, we describe a modification of the line to include a constitutive mCherry reporter integrated into the SH4-2 safe harbor locus, enabling fluorescent tracking of microglia in mixed cultures or in vivo. This protocol provides a reproducible and scalable platform for generating functional human microglia from a widely used hiPSC line, supporting applications in disease modeling, neuroinflammation research, and therapeutic screening.

Journal Article↗
PubMed2025

Heterochromatin-based silencing of a foreign tandem repeat in Drosophila melanogaster shows unusual biochemistry and temperature sensitivity.

Eukaryotic genomes are packaged into chromatin, a regulatory nucleoprotein assembly. Establishment, maintenance, and interconversion of chromatin states is required for correct patterns of gene expression, genome integrity, and survival. Transcriptionally repressive heterochromatin minimizes mobilization of transposable elements and limits expansion of other repetitive DNA, but mechanisms for recognition of the latter sequences are not well established. We previously demonstrated in Drosophila melanogaster that transcripts derived from 1360 and Invader4 transposon insertions can trigger local conversion of transcriptionally permissive euchromatin to heterochromatin through the piRNA system, but only in a subset of genomic locations near existing blocks of heterochromatin. Here we show that a ~9 kb tandem array of the 36-nucleotide lac operator (lacO) sequence of Escherichia coli can form ectopic heterochromatin at a similar subset of sites, resulting in variegating expression of an adjacent reporter gene. Heterochromatin Protein 1a (HP1a) and histone deacetylation are required for lacO repeat-induced silencing, but, contrasting with previously described Position Effect Variegation (PEV), we do not observe increased histone H3 lysine 9 methylation. Silencing is effective at 25°C and suppressed at 18°C (in contrast to canonical PEV, which is enhanced at 18°C), indicating involvement of a temperature-sensitive component. Temperature switching experiments show that lacO repeat-induced heterochromatin formation is reversible throughout larval development following an HP1a-dependent initiation step in the early embryo. We conclude that the Drosophila nucleus can recognize a completely foreign tandem repeat as a target for heterochromatin formation, and that the heterochromatin structure established is distinct from that of endogenous tandem arrays.

HP1a↗
PubMed2025

Polygenic Risk Scores and HLA Class II Variants are Biomarkers of Corticosteroid Response in Childhood Nephrotic Syndrome.

INTRODUCTION: Nephrotic syndrome (NS), a common glomerular disease in children, is classified based on response to corticosteroid therapy as either steroid-sensitive nephrotic syndrome (SSNS), or steroid-resistant nephrotic syndrome (SRNS). However, there are currently no reliable predictors of therapy response at initial clinical presentation. METHODS: We conducted genome-wide association studies, developed polygenic risk scores (PRS) for therapy response and analyzed classical HLA alleles in 1,997 (994 discovery and 1,003 replication/validation cohorts) previously unstudied children with NS and 3,558 ancestry-matched controls. RESULTS: A significant association with HLA loci defined by variants in HLA-DQB1, HLA-DRB1, and HLA-DQA1 were found for SSNS (but not SRNS), along with a second immune-related SSNS locus: CLEC16A. A PRS that discriminates between SSNS and SRNS was validated in two independent cohorts. The HLA haplotype HLA- DRB1*07:01~DQA1*02:01~DQB1*02:02 was associated with ~4 times the risk of developing SSNS. A model incorporating HLA haplotype, PRS score, and age at onset of the disease was the best predictor of steroid responsiveness with an AUC of 0.68-0.70 and an overall classification accuracy of SSNS versus SRNS of 67-71%. CONCLUSIONS: Our findings confirm that SSNS (unlike SRNS) is an immune-mediated HLA-associated disorder. The PRS for therapy response and HLA haplotype can serve as biomarkers and provide a foundation for more accurate diagnoses and tailored and individualized treatment.

HLA haplotype↗
PubMed2025

A Sequence Motif Enables Widespread Use of Non-Canonical Redox Cofactors in Natural Enzymes.

Non-canonical redox cofactors (NRCs) are promising alternatives to nicotinamide adenine dinucleotide (phosphate) (NAD(P)+) for biomanufacturing due to low cost and exquisite electron delivery control, yet their adoption is limited by the scarcity of compatible enzymes. Here, we screened the aldehyde dehydrogenase (ALDH) protein family and identified a conserved RH/QxxR sequence motif that enables widespread NRC activity among natural enzymes. Bos taurus ALDH3a1 and Pseudanabaena biceps ALDH exhibit unprecedented turnover with nicotinamide mononucleotide (NMN+), with kcat values matching or exceeding that of NAD+ and surpassing most engineered NRC-active enzymes by 10 to 105-fold, based on the relative NRC to native activity. Structural and dynamic analyses reveal this motif reinforces cofactor positioning and pre-organizes the active site without dependence on the adenosine monophosphate moiety of NAD+. When introduced into diverse ALDH scaffolds, the RH/QxxR motif enhances NMN+ activity up to 60-fold. In addition to NMN+, this motif also supports activity across multiple non-nucleotide, simple synthetic NRCs such as 1-(2-carbamoylmethyl)nicotinamide (AmNA+). These findings elucidate Nature's solution to the engineering challenge of obtaining NRC-active enzymes and offers a blueprint to mine latent evolutionary plasticity in natural enzymes that serve as superior engineering starting points.

Active site pre-organization↗