Biology subjects

Smith, K. M.

Publications and source records attributed to Smith, K. M..

18 recordsLinked to original sources

bioRxiv2026

Polyketide synthase 12 is an in vivo essential source of novel mycolyl lipids in Mycobacterium tuberculosis

Mycobacterium tuberculosis (Mtb) is a major pathogen worldwide that infects and transmits only among humans, yet nearly all in vivo virulence research occurs in non-human hosts. To overcome this central challenge in tuberculosis research, we leveraged a dataset of more than 50,000 sequenced isolates to identify Mtb genes that that are functionally preserved during natural infection, disease causation, and transmission between humans. This whole-genome ranking identified polyketide synthase 12 (pks12 ) as an exceptionally in vivo essential gene in human tuberculosis, which we validated in zebrafish and mouse models. Though Pks12 produces a mycoketide lipid in only trace amounts, pks12 deletion severely altered the host-facing surface and arabinoglycan architecture of Mtb. This amplified effect was explained through the discovery of mannosyl-{beta}-1-phosphomycoketide monomycolate (MPMMM), which is synthesized from the known Pks12 product at higher abundance by antigen 85 mycolyltransferases. Thus, we used a new host-facing genomic-metabolomic-phenotypic approach to discover the functions of an in vivo essential Mtb gene, which controls the physical structure of the Mtb-host interface.

microbiology↗

bioRxiv2025

NGN2 Expression and Regional Patterning Allow Rapid Differentiation from hiPSCs to DRG-Like Neurons Responsive to Type 2 Cytokines

Itch or pruritus, is a sensation that elicits scratching behaviour and is a major symptom and cause of morbidity in skin diseases such as atopic dermatitis (AD), allergic contact dermatitis (ACD), prurigo nodularis (PN), and urticaria. Itch is often triggered by inflammatory stimuli in the skin including type 2 cytokines such as IL-4, IL-13, and/or IL-31. Several therapies targeting type 2 immune pathways have been developed to treat pruritus; however, itch improvement in many patients remains to be improved. Thus, additional approaches to modulate sensory neuron activity are needed. Ex vivo or even in vitro study of the molecular mechanisms underlying primary sensory neuron activation is challenging since harvesting neurons from dorsal root ganglia (DRG) in patients can only be done from cadavers. Herein, we describe rapid human sensory neurons generation (2 days to precursor cells) by in vitro differentiation of human induced pluripotent stem cells (hiPSC) from simultaneous application of patterning factors with NGN2 overexpression. We show that these hiPSC-derived sensory neurons possess key characteristics of primary sensory neurons. They express key neuronal markers, such as TRKA receptors, TRPV1 and TRPA1 channels, and functionally respond to the TRPV1 agonist capsaicin. In addition, they express key type 2 cytokine receptors such as interleukin (IL)-4R and IL31-R, known to promote itch in AD and PN. Moreover, these cells are functional as our sensory neurons respond to IL-4, IL-13 and IL-31 stimulation. Collectively, these data demonstrate that our protocol generates a phenotypic profile consistent with native somatosensory neurons that can facilitate development of novel approaches to model and treat pruritic disease.

neuroscience↗

bioRxiv2025

Transcriptomic profiling and machine learning uncover gene signatures of psoriasis endotypes and disease severity

BackgroundDespite increased understanding of psoriasis pathogenesis, molecular classification of clinical phenotypes and disease severity is poorly defined. Knowledge gaps include whether molecular endotypes of psoriasis underlie distinct clinical phenotypes and the positive and negative molecular regulators of disease severity across tissue compartments. MethodsWe performed comprehensive RNA-sequencing of skin and blood (n=718) from prospectively-recruited, deeply-phenotyped discovery and replication cohorts of 146 subjects with moderate-to-severe psoriasis initiating TNF-inhibitor (adalimumab) or IL-12/23-inhibitor (ustekinumab) therapy. ResultsUsing two complementary methods for dimensionality reduction, we defined distinct but interconnected co-expression modules and factors within skin and blood that were significantly associated with disease phenotypes and disease severity, as measured by Psoriasis Area Severity Index (PASI). We identified a 14-gene signature negatively associated with BMI in nonlesional skin and disease severity in lesional skin, respectively. Genotype integration revealed that HLA-DQA1*01 and HLA-DRB1*15 genotypes were positively associated with baseline disease severity. Using Gaussian process regression followed by SHAP (SHapley Additive exPlanations), we defined two core drug independent and disease severity-associated gene modules in lesional skin - one positive, one negative - and a lesional 9-gene signature predictive of disease severity. Disease severity signatures in blood were only seen following adalimumab exposure, suggesting greater systemic impact of adalimumab compared to ustekinumab, in line with its side effect profile. In contrast, a gene signature in blood linked to HLA-C*06:02 status was independent of disease severity or drug. ConclusionsThese findings delineate gene-environmental and genetic effects on the psoriasis transcriptome linked to disease severity. Plain Language SummaryPsoriasis is a common and debilitating skin disease, linked to multiple other inflammatory conditions. A lot is known about the mechanism of psoriasis and its inherited and external influences. Despite this, doctors cannot yet offer personalised treatments as it has been difficult to discover whether biological pathways are associated with disease severity, response to treatment or a persons likelihood of having other linked diseases. To help address this, we collected skin and blood samples and the personal characteristics of a group of people with severe psoriasis across the United Kingdom. Using computer-based methods, we discovered common biological processes underlying different psoriasis types, including genes that connect psoriasis severity with obesity, and another set of genes that help predict disease severity.

systems biology↗

bioRxiv2025

Induction of Human Pruriceptors from Pluripotent Stem Cells via Transcription Factors

Pruriception is crucial for defense against external stimuli but can lead to chronic pruritus, a debilitating condition affecting millions worldwide. Our understanding of the cellular and molecular mechanisms behind the sensation of itch has been hindered by the lack of functional human models. Here, we address this limitation by developing a protocol to generate induced pruriceptors (iPruriceptors) from human pluripotent stem cells (hPSCs). We compared two differentiation approaches: a direct method via forced expression of transcription factors (TFs) in hPSCs, and a 2-step process through expression of TFs in hPSC-derived neural crest cells (NCCs). The 2-step protocol proved superior in inducing a transcriptional program that closely resembles that of human pruriceptors. Our optimized protocol employs forced expression of NGN1 and ISL1 to drive differentiation from NCCs into pruriceptors, enhancing the expression of known pruritogen receptors such as IL31RA, which pairs with OSMR, and HRH1. The induction of this transcriptional program leads to functional maturation of iPruriceptors. Accordingly, iPruriceptors exhibit robust responses to itch stimuli and in vivo-like itch pharmacology such as treatment with ABT-317, a JAK1 inhibitor tool compound, similar to those targeting intensive pruritus in atopic dermatitis (AD). Importantly, iPruriceptors can be generated without viral vectors or safe-harbor gene editing, using a PiggyBac-based transfection method that simplifies scalability. Our protocol offers a robust platform for investigating itch biology, modeling chronic pruritus, and enabling high-throughput screening for therapeutic target discovery. HighlightsO_LINGN1 and ISL1 forced expression in NCCs induces rapid differentiation to iPruriceptors C_LIO_LIiPruriceptors share transcriptional profile of primary human pruriceptors C_LIO_LIiPruriceptors have electrophysiological responses to known pruritogens C_LIO_LIiPruriceptors have JAK1-dependent IL-31/-13 responses blocked by ABT-317 C_LI

neuroscience↗

bioRxiv2025

Relative strength variability measures for brain structural connectomes and their relationship with cognitive functioning

In this work, we propose a new class of graph measures for weighted connectivity information in the human brain based on node relative strengths: relative strength variability (RSV), measuring susceptibility to targeted attacks, and hierarchical RSV (hRSV), a first weighted statistical complexity measure for networks. Using six different network weights for structural connectomes from the UK Biobank, we conduct comprehensive analyses to explore relationships between the RSV and hRSV, and (i) other known network measures, (ii) general cognitive function ( g). Both measures exhibit low correlations with other graph measures across all connectivity weightings indicating that they capture new information of the brain connectome. We found higher g was associated with lower RSV and lower hRSV. That is, higher g was associated with higher resistance to targeted attack and lower statistical complexity. Moreover, the proposed measures had consistently stronger associations with g than other widely used graph measures including clustering coefficient and global efficiency and were incrementally significant for predicting g above other measures for five of the six network weights. Overall, we present a new class of weighted network measures based on variations of relative node strengths which significantly improved prediction of general cognition from traditional weighted structural connectomes.

neuroscience↗

bioRxiv2025

Learning robust gene expression embeddings for disease subtyping, biomarker identification and cross-species alignment

High dimension gene expression measurements such as microarray and RNA-seq data, are often plagued by sources of unwanted variation. This variability can lead to the obscuring of meaningful biological signal by technical noise and non-interesting biological variation, thus resulting in failure to identify the same set of targets and biomarkers in independent studies. This phenomenon contributes to the so-called reproducibility crisis and makes preclinical drug development more challenging. There is an urgent need for an improved process to identify shared biological signals in large datasets from independent cohorts. In this article, we propose an innovative method called Joint-Embedding via Canonical Correlation Analysis (JECCA), which aims to capture relevant biological variation by learning of shared lower dimension embedding across multiple datasets. We demonstrate the efficacy of JECCA in analyzing multiple human cancer and inflammatory disease gene expression datasets, showcasing its ability to identify robust, biologically relevant signals. By leveraging these embeddings, our approach enables several important applications: (1) robust disease endotype identification by joint unsupervised analysis of multiple independent gene expression datasets (2) reproducible and biologically meaningful biomarker identification to enhance prediction of treatment outcome (3) cross-platform analysis, such as integration of RNA-seq and microarray data, which are accumulating exponentially in public databases (4) cross-species analysis to identify conserved molecular features between human and mouse. We provide comparative analyses that highlight the superior performance of JECCA over other approaches such as batch value-corrected methods based on joint analysis. As analysis of publicly available datasets becomes more prevalent, we foresee JECCA being employed in multiple disease contexts.

bioinformatics↗

bioRxiv2023

Identification of a convergent spinal neuron population that encodes itch

Itch is a protective sensation that drives scratching. Although specific cell types have been proposed to underlie itch, the neural circuit basis for itch remains unclear. Here, we used two-photon Ca2+ imaging of the dorsal horn to visualize the neuronal populations that are activated by itch-inducing agents. We identify a convergent population of spinal neurons that is defined by the expression of GRPR. Moreover, we discover that itch is conveyed to the brain via GRPR-expressing spinal output neurons that target the lateral parabrachial nucleus. Further, we show that nalfurafine, a clinically effective kappa opioid receptor agonist, relieves itch by inhibiting GRPR spinoparabrachial neurons. Finally, we demonstrate that a subset of GRPR spinal neurons show persistent, cell-intrinsic Ca2+ oscillations. These experiments provide the first population-level view of the spinal neurons that respond to pruritic stimuli, pinpoint the output neurons that convey itch to the brain, and identify the cellular target of kappa opioid receptor agonists for the inhibition of itch. In briefThrough population imaging, Sheahan et al. identify a network of neurons in the dorsal horn that is activated by pruritogens and find that kappa opioid receptor signaling inhibits itch through the selective inhibition of GRPR spinoparabrachial neurons. HighlightsO_LIItch-inducing agents drive activity in a common population of GRPR-expressing spinal interneurons C_LIO_LIGRPR spinal projection neurons transmit itch from the spinal cord to the brain C_LIO_LIKappa opioids reduce itch through the inhibition of GRPR spinoparabrachial neurons C_LIO_LIGRPR activation elicits persistent, intrinsic Ca2+ oscillations C_LI

neuroscience↗

bioRxiv2023

Calretinin-expressing islet cells: a source of pre- and post-synaptic inhibition of non-peptidergic nociceptor input to the mouse spinal cord

Unmyelinated non-peptidergic nociceptors (NP afferents) arborise in lamina II of the spinal cord and receive GABAergic axoaxonic synapses, which mediate presynaptic inhibition. However, until now the source of this axoaxonic synaptic input was not known. Here we provide evidence that it originates from a population of inhibitory calretinin-expressing interneurons (iCRs), which correspond to lamina II islet cells. The NP afferents can be assigned to 3 functionally distinct classes (NP1-3). NP1 afferents have been implicated in pathological pain states, while NP2 and NP3 afferents also function as pruritoceptors. Our findings suggest that all 3 of these afferent types innervate iCRs and receive axoaxonic synapses from them, providing feedback inhibition of NP input. The iCRs also form axodendritic synapses, and their targets include cells that are themselves innervated by the NP afferents, thus allowing for feedforward inhibition. The iCRs are therefore ideally placed to control the input from non-peptidergic nociceptors and pruritoceptors to other dorsal horn neurons, and thus represent a potential therapeutic target for the treatment of chronic pain and itch.

neuroscience↗

bioRxiv2022

The delta-opioid receptor bidirectionally modulates itch

Opioid signaling has been shown to be critically important in the neuromodulation of sensory circuits in the superficial spinal cord. Agonists of the mu-opioid receptor (MOR) elicit itch, whereas agonists of the kappa-opioid receptor (KOR) have been shown to inhibit itch. Despite the clear roles of MOR and KOR for the modulation itch, whether the delta-opioid receptor (DOR) is involved in the regulation of itch remained unknown. Here, we show that intrathecal administration of DOR agonists suppresses chemical itch and that intrathecal application of DOR antagonists is sufficient to evoke itch. We identify that spinal enkephalin neurons co-express neuropeptide Y (NPY), a peptide previously implicated in the inhibition of itch. In the spinal cord, DOR overlapped with both the NPY receptor (NPY1R) and KOR, suggesting that DOR neurons represent a site for convergent itch information in the dorsal horn. Lastly, we found that neurons co-expressing DOR and KOR showed significant Fos induction following pruritogen-evoked itch. These results uncover a role for DOR in the modulation of itch in the superficial dorsal horn. PerspectiveThis article reveals the role of the delta opioid receptor in itch. Intrathecal administration of delta agonists suppresses itch whereas the administration of delta antagonists is sufficient to induce itch. These studies highlight the importance of delta-opioid signaling for the modulation of itch behaviors, which may represent new targets for the management of itch disorders.

neuroscience↗

bioRxiv2021

Early life sleep disruption drives lasting sex-specific changes in behavior in genetically vulnerable Shank3 heterozygous autism model mice

BackgroundPatients with autism spectrum disorder (ASD) experience high rates of sleep disruption beginning early in life, however the developmental consequences of this disruption are not understood. We examined sleep behavior and the consequences of sleep disruption in developing mice bearing C-terminal truncation mutation in the high-confidence ASD risk gene SHANK3 (Shank3{Delta}C). We hypothesized that sleep disruption may be an early sign of developmental divergence, and that clinically-relevant Shank3WT/{Delta}C mice may be at increased risk of lasting deleterious outcomes following early-life sleep disruption. MethodsWe recorded sleep behavior in developing Shank3{Delta}C/{Delta}C, Shank3WT/{Delta}C, and wild type siblings of both sexes using a non-invasive home cage monitoring system. Separately, litters of Shank3WT/{Delta}C and wildtype littermates were exposed to automated mechanical sleep disruption for 7 days prior to weaning (early-life sleep disruption: ELSD) or post-adolescence (PASD) or undisturbed control (CON) conditions. All groups underwent standard behavioral testing as adults. ResultsMale and female Shank3{Delta}C/{Delta}C mice slept significantly less than wild type and Shank3WT/{Delta}C siblings shortly after weaning, with increasing sleep fragmentation in adolescence, indicating that sleep disruption has a developmental onset in this ASD model. ELSD treatment interacted with genetic vulnerability in Shank3WT/{Delta}Cmice, resulting in lasting, sex-specific changes in behavior, whereas wildtype siblings were largely resilient to these effects. Male ELSD Shank3WT/{Delta}Csubjects demonstrated significant changes in sociability, sensory processing, and locomotion, while female ELSD Shank3WT/{Delta}C subjects had a significant reduction in risk aversion. CON Shank3WT/{Delta}C mice, PASD mice, and all wildtype mice demonstrated typical behavioral responses in most tests. LimitationsThis study tested the interaction between developmental sleep disruption and genetic vulnerability using a single ASD mouse model: Shank3{Delta}C (deletion of exon 21). The broader implications of this work should be supported by additional studies using ASD model mice with distinct genetic vulnerabilities. ConclusionOur study shows that sleep disruption during sensitive periods of early life interact with underlying genetic vulnerability to drive lasting and sex-specific changes in behavior. As individuals progress through maturation they gain resilience to the lasting effects of sleep disruption. This work highlights developmental sleep disruption as an important vulnerability in ASD susceptibility.

animal behavior and cognition↗

bioRxiv2021

Progesterone receptor-A isoform interaction with RUNX transcription factors controls chromatin remodelling at promoters during ovulation

Progesterone receptor (PGR) plays diverse roles in reproductive tissues and thus coordinates mammalian fertility. In the ovary, acutely induced PGR is the key determinant of ovulation through transcriptional control of a unique set of genes that culminates in follicle rupture. However, the molecular mechanisms for PGRs specialised function in ovulation is poorly understood. To address this, we assembled a detailed genomic profile of PGR action through combined ATAC-seq, RNA-seq and ChIP-seq analysis in wildtype and isoform-specific PGR null mice. We demonstrated the unique action of PGR-A isoform in the ovary through a transcriptional complex involving physical interaction with RUNX and JUN/FOS transcription factors. The assembly of this unique complex directs targeting of PGR binding to proximal promoter regions and enables chromatin accessibility, leading to ovulatory gene induction. This PGR signalling mechanism is specific to ovulation and provides potential targets for infertility treatments as well as new contraceptives that block ovulation.

molecular biology↗

bioRxiv2020

Three neurotransmitters regulate diverse inhibitory and excitatory Parvalbumin interneuron circuits in the dorsal horn

Parvalbumin-expressing interneurons (PVINs) in the spinal dorsal horn are found primarily in laminae II inner and III. Inhibitory PVINs (iPVINs) play an important in segregating innocuous tactile input from pain-processing circuits, achieved through presynaptic inhibition of myelinated low-threshold mechanoreceptors and postsynaptic inhibition of distinct spinal circuits. By comparison, relatively little is known of the role of excitatory PVINs (ePVINs) in sensory processing. Here we use neuroanatomical and optogenetic approaches to show that ePVINs comprise a larger proportion of the PVIN population than previously reported, and that both ePVIN and iPVIN populations form synaptic connections amongst (and between) themselves. We find that these cells contribute to neuronal networks that influence activity within several functionally distinct circuits, and that aberrant activity of ePVINs under pathological conditions contributes to the development of mechanical hypersensitivity.

neuroscience↗

bioRxiv2020

Optogenetic evaluation of the ability of different cutaneous C-fiber afferents to evoke aversive behaviors

Most cutaneous C-fibers, including both peptidergic and non-peptidergic subtypes are presumed to be nociceptors and respond to noxious input in a graded manner. However, mechanically sensitive, non-peptidergic C-fibers also respond to mechanical input in the innocuous range, and so the degree to which they contribute to nociception remains unclear. To address this gap, we investigated the function of non-peptidergic afferents using the MrgprdCre allele. In real time place aversion studies, we found that low frequency optogenetic activation of MrgrpdCre lineage neurons was not aversive in naive mice, but became aversive after spared nerve injury (SNI). To address the underlying mechanisms of this allodynia, we recorded from lamina I spinoparabrachial (SPB) neurons using the semi-intact ex vivo preparation. Following SNI, innocuous brushing of the skin gave rise to abnormal activity in lamina I SPB neurons, consisting of an increase in the proportion of recorded neurons that responded with excitatory post synaptic potentials or action potentials. This increase was likely due, at least in part, to an increase in the proportion of lamina I (LI) SPB neurons that received input upon optogenetic activation of MrgprdCre lineage neurons. Intriguingly, in SPB neurons there was a significant increase in the EPSC latency from MrgprdCre lineage input following SNI, consistent with the possibility that the greater activation post SNI could be due to the recruitment of a new polysynaptic circuit. Together, our findings suggest MrgprdCre lineage neurons can provide mechanical input to the dorsal horn that is non-noxious before injury but becomes noxious afterwards due the engagement of a previously silent polysynaptic circuit in the dorsal horn.

neuroscience↗

bioRxiv2020

Resilience and evolvability of protein-protein interaction networks

Protein-protein interaction (PPI) networks represent complex intra-cellular protein interactions, and the presence or absence of such interactions can lead to biological changes in an organism. Recent network-based approaches have shown that a phenotypes PPI networks resilience to environmental perturbations is related to its placement in the tree of life; though we still do not know how or why certain intra-cellular factors can bring about this resilience. Here, we explore the influence of gene expression and network properties on PPI networks resilience. We use publicly-available data of PPIs for E. coli, S. cerevisiae, and H. sapiens, where we compute changes in network resilience as new nodes (proteins) are added to the networks under three node addition mechanisms--random, degree-based, and gene-expression-based attachments. By calculating the resilience of the resulting networks, we estimate the effectiveness of these node addition mechanisms. We demonstrate that adding nodes with gene-expression-based preferential attachment (as opposed to random or degree-based) preserves and can increase the original resilience of PPI network in all three species, regardless of gene expression distribution or network structure. These findings introduce a general notion of prospective resilience, which highlights the key role of network structures in understanding the evolvability of phenotypic traits.

systems biology↗

bioRxiv2019

United States wildlife and wildlife product imports from 2000-2014

The global wildlife trade network is a massive system that has been shown to threaten biodiversity conservation, introduce non-native species and pathogens, and cause chronic animal welfare concerns. Despite its scale and impact, comprehensive characterization of the global wildlife trade is hampered by data that are limited in their temporal or taxonomic scope and detail. To help fill this gap, we present data on 15 years of the importation of wildlife and their derived products into the United States (2000-2014), originally collected by the United States Fish and Wildlife Service. We curated and cleaned the data and added taxonomic information to improve data usability. These data include > 2 million wildlife or wildlife product shipments, representing > 60 biological classes and > 3.2 billion live organisms. These data will be broadly useful to both scientists and policymakers seeking to better understand the volume, sources, biological composition, and potential risks of the global wildlife trade.

ecology↗

bioRxiv2019

Calretinin positive neurons form an excitatory amplifier network in the spinal cord dorsal horn

The passage of nociceptive information is relayed through the spinal cord dorsal horn, a critical area in sensory processing. The neuronal circuits in this region that underpin sensory perception must be clarified to better understand how dysfunction can lead to pathological pain. This study used an optogenetic approach to selectively activate neurons that contain the calcium-binding protein calretinin (CR). We show that CR+ interneurons form an interconnected network that can initiate and sustain enhanced excitatory signaling, and directly relays signals to lamina I projection neurons. In vivo photoactivation of CR+ interneurons resulted in a significant nocifensive behavior that was morphine sensitive and cause a conditioned place aversion. Furthermore, halorhodopsin-mediated inhibition of CR+ interneurons elevated sensory thresholds. These results suggest that neuronal circuits in the superficial dorsal horn that involve excitatory CR+ neurons are important for the generation and amplification of pain, and identify these interneurons as a future analgesic target.

neuroscience↗

bioRxiv2018

The multi-speed genome of Fusarium oxysporum reveals association of histone modifications with sequence divergence and footprints of past horizontal chromosome transfer events

Fusarium oxysporum is an economically important pathogen causing wilting or rotting disease symptoms in a large number of crops. It is proposed to have a structured, \"two-speed\" genome: i.e. regions containing genes involved in pathogenicity cluster with transposons on separate accessory chromosomes. This is hypothesized to enhance evolvability. Given the continuum of adaptation of all the genes encoded in a genome, however, one would expect a more complex genome structure. By comparing the genome of reference strain Fol4287 to those of 58 other Fusarium oxysporum strains, we found that some Fol4287 accessory chromosomes are lineage-specific, while others occur in multiple lineages with very high sequence similarity - but only in strains that infect the same host as Fol4287. This indicates that horizontal chromosome transfer has been instrumental in past host-switches. Unexpectedly, we found that the sequence of the three smallest core chromosomes (Chr. 11, 12 and 13) is more divergent than that of the other core chromosomes. Moreover, these chromosomes are enriched in genes involved in metabolism and transport and genes that are differentially regulated during infection. Interestingly, these chromosomes are -like the accessory chromosomes- marked by histone H3 lysine 27 trimethylation (H3K27me3) and depleted in histone H3 lysine 4 dimethylation (H3K4me2). Detailed genomic analyses revealed a complex, \"multi-speed genome\" structure in Fusarium oxysporum. We found a strong association of H3K27me3 with elevated levels of sequence divergence that is independent of the presence of repetitive elements. This provides new leads into how clustering of genes evolving at similar rates could increase evolvability.\n\nAuthor summaryFungi that cause disease on plants are an increasingly important threat to food security. New fungal diseases emerge regularly. The agricultural industry makes large investments to breed crops that are resistant to fungal infections, yet rapid adaptation enables fungal pathogens to overcome this resistance within a few years or decades. It has been proposed that genome compartmentalization of plant pathogenic fungi, in which infection-related genes are clustered with transposable elements (or jumping genes) into separate, fast-evolving regions, enhances their adaptivity. Here, we aimed to shed light on the possible interplay between genome organization and adaptation. We measured differences in sequence divergence and dispensability between and within individual chromosomes of the important plant pathogen Fusarium oxysporum. Based on these differences we defined four distinct chromosomal categories. We then mapped histone modifications and gene expression levels under different conditions for these four categories. We found a division of labor between chromosomes, where some are pathogenicity chromosomes - specialized towards infection of a specific host, while others are enriched in genes involved in more generic infection-related processes. Moreover, we confirmed that horizontal transfer of pathogenicity chromosomes likely plays an important role in gain of pathogenicity. Finally, we found that a specific histone modification is associated with increased sequence divergence.

genomics↗

bioRxiv2018

Destabilization of chromosome structure by histone H3 lysine 27 methylation

Chromosome and genome stability are important for normal cell function as instability often correlates with disease and dysfunction of DNA repair mechanisms. Many organisms maintain supernumerary or accessory chromosomes that deviate from standard chromosomes. The pathogenic fungus Zymoseptoria tritici has as many as eight accessory chromosomes, which are highly unstable during meiosis and mitosis, transcriptionally repressed, show enrichment of repetitive elements, and enrichment with heterochromatic histone methylation marks, e.g., trimethylation of H3 lysine 9 or lysine 27 (H3K9me3, H3K27me3). To elucidate the role of heterochromatin on genome stability in Z. tritici, we deleted the genes encoding the methyltransferases responsible for H3K9me3 and H3K27me3, kmt1 and kmt6, respectively, and generated a double mutant. We combined experimental evolution and genomic analyses to determine the impact of these deletions on chromosome and genome stability, both in vitro and in planta. We used whole genome sequencing, ChIP-seq, and RNA-seq to compare changes in genome and chromatin structure, and differences in gene expression between mutant and wildtype strains. Analyses of genome and ChIP-seq data in H3K9me3-deficient strains revealed dramatic chromatin reorganization, where H3K27me3 is mostly relocalized into regions that are enriched with H3K9me3 in wild type. Many genome rearrangements and formation of new chromosomes were found in the absence of H3K9me3, accompanied by activation of transposable elements. In stark contrast, loss of H3K27me3 actually increased the stability of accessory chromosomes under normal growth conditions in vitro, even without large scale changes in gene activity. We conclude that H3K9me3 is important for the maintenance of genome stability because it disallows H3K27me3 in these regions. In this system, H3K27me3 reduces the overall stability of accessory chromosomes, generating a \"metastable\" state for these quasi-essential regions of the genome.\n\nAuthor SummaryGenome and chromosome stability are essential to maintain normal cell function and viability. However, differences in genome and chromosome structure are frequently found in organisms that undergo rapid adaptation to changing environmental conditions, and in humans are often found in cancer cells. We study genome instability in a fungal pathogen that exhibits a high degree of genetic diversity. Regions that show extraordinary diversity in this pathogen are the transposon-rich accessory chromosomes, which contain few genes that are of unknown benefit to the organism but maintained in the population and thus considered \"quasi essential\". Accessory chromosomes in all fungi studied so far are enriched with markers for heterochromatin, namely trimethylation of H3 lysine 9 and 27 (H3K9me3, H3K27me3). We show that loss of these heterochromatin marks has strong but opposing effects on genome stability. While loss of the transposon-associated mark H3K9me3 destabilizes the entire genome, presence of H3K27me3 favors instability of accessory chromosomes. Our study provides insight into the relationship between chromatin and genome stability and why some regions are more susceptible to genetic diversity than others.

genetics↗