Biology subjects

Parker, J.

Publications and source records attributed to Parker, J..

50 records · Page 3Linked to original sources

bioRxiv2020

Aberrant gliogenesis and excitation in MEF2C autism patient hiPSC-neurons and cerebral organoids

MEF2C has been shown to be a critical transcription factor for neurodevelopment, whose loss-of-function mutation in humans results in MEF2C haploinsufficiency syndrome (MHS), a severe form of autism spectrum disorder (ASD)/intellectual disability (ID). Here, we use patient hiPSC-derived cerebrocortical neurons and cerebral organoids to characterize MHS deficits. Unexpectedly, we found an aberrant micro-RNA-mediated gliogenesis pathway that contributes to decreased neurogenesis. We also demonstrate network-level hyperexcitability in neurons, as evidenced by excessive synaptic and extrasynaptic activity contributing to excitatory/inhibitory (E/I) imbalance. Notably, the extrasynaptic NMDA receptor antagonist, NitroSynapsin, corrects this aberrant electrical activity associated with abnormal phenotypes. During neurodevelopment, MEF2C regulates many ASD-associated gene networks suggesting that our approach may lead to personalized therapy for multiple forms of ASD. One sentence summaryAutism-like MEF2C+/- patient hiPSC models show miRNA-mediated overproduction of astrocytes and hyperactivity of neurons.

neuroscience↗

bioRxiv2020

YY1 cistrome analysis uncovers an essential requirement of the YY1:BRD4-PFKP regulatory axis for promoting tumorigenesis of castration-resistant prostate cancer

Castration-resistant prostate cancer (CRPC) is a terminal disease, demanding a better understanding of its pathogenesis. Targeted therapy needs to be developed for CRPC due to its heterogeneity and resistance to current treatments. Here, through cistrome study of YY1, a transcription factor significantly overexpressed during prostate cancer progression, we identify a YY1-PFKP axis to be essential for CRPC tumorigenesis. Depletion of YY1 in independent CRPC models dramatically reduced tumor cell growth in vitro and delayed oncogenic progression in vivo. Importantly, YY1 functions as a master regulator of prostate tumor metabolism including the Warburg effect and mitochondria respiration. Loss-of-function and rescue studies further reveals a mechanistic underpinning in which YY1 directly binds and trans-activates PFKP, a gene encoding the rate-limiting enzyme for glycolysis, significantly contributing to the YY1-enforced oncogenic phenotypes such as enhanced tumor cell glycolysis and malignant growth. Additionally, a vast majority of gene-regulatory element in advanced prostate cancer cells are bound by YY1, lending a support for its role as a master regulator of prostate cancer progression. YY1 interactome studies point to bromodomain-containing coactivators in prostate cancer, which act as functional partners of YY1 to potentiate YY1-related target gene activation. Altogether, this study unveils an unexplored YY1:BRD4-PFKP oncogenic axis operating in advanced prostate cancer with implications for therapy.

cancer biology↗

bioRxiv2020

Mapping the SARS-CoV-2 spike glycoprotein-derived peptidome presented by HLA class II on dendritic cells

Understanding and eliciting protective immune responses to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an urgent priority. To facilitate these objectives, we have profiled the repertoire of human leukocyte antigen class II (HLA-II)-bound peptides presented by HLA-DR diverse monocyte-derived dendritic cells pulsed with SARS-CoV-2 spike (S) protein. We identify 209 unique HLA-II-bound peptide sequences, many forming nested sets, which map to sites throughout S including glycosylated regions. Comparison of the glycosylation profile of the S protein to that of the HLA-II-bound S peptides revealed substantial trimming of glycan residues on the latter, likely introduced during antigen processing. Our data also highlight the receptor-binding motif in S1 as a HLA-DR-binding peptide-rich region. Results from this study have application in vaccine design, and will aid analysis of CD4+ T cell responses in infected individuals and vaccine recipients.

immunology↗

bioRxiv2020

Rapid Parallel Adaptation to Anthropogenic Heavy Metal Pollution

The impact of human mediated environmental change on the evolutionary trajectories of wild organisms is poorly understood. In particular, species capacity to adapt rapidly (in hundreds of generations or less), reproducibly and predictably to extreme environmental change is unclear. Silene uniflora is predominantly a coastal species, but it has also colonised isolated, disused mines with phytotoxic, zinc-contaminated soils. Here, we found that rapid parallel adaptation to anthropogenic pollution has taken place without geneflow spreading adaptive alleles between populations of the mine ecotype. Across replicate ecotype pairs, we identified shared targets of selection with functions linked to physiological differences between the ecotypes, although the genetic response is only partially shared between mine populations. Our results are consistent with a complex, polygenic genetic architecture underpinning rapid adaptation. This shows that even under a scenario of strong selection and rapid adaptation, evolutionary responses to human activities may be idiosyncratic at the genetic level and, therefore, difficult to predict from genomic data.

evolutionary biology↗

bioRxiv2020

Epigenomic characterization of latent HIV infection identifies latency regulating transcription factors.

Transcriptional silencing of HIV generates a reservoir of latently infected cells, but the mechanisms that lead to this outcome are not well understood. We characterized a primary cell model of HIV latency, and observed that latency is a stable, heritable viral state that is rapidly reestablished after stimulation. Using Assay of Transposon-Accessible Chromatin sequencing (ATACseq) we found that latently infected cells exhibit reduced proviral accessibility, elevated activity of Forkead and Kruppel-like factor transcription factors (TFs), and reduced activity of AP-1, RUNX and GATA TFs. Latency reversing agents caused distinct patterns of chromatin reopening across the provirus. Furthermore, depletion of a chromatin domain insulator, CTCF inhibited HIV latency, identifying this factor as playing a key role in the initiation or enforcement of latency. These data indicate that HIV latency develops preferentially in cells with a distinct pattern of TF activity that promotes a closed proviral structure and inhibits viral gene expression.

microbiology↗

bioRxiv2020

Reovirus σ3 protein limits interferon expression and cell death induction

Induction of necroptosis by mammalian reovirus requires both type I interferon (IFN)-signaling and viral replication events that lead to production of progeny genomic dsRNA. The reovirus outer capsid protein {micro}1 negatively regulates reovirus-induced necroptosis by limiting RNA synthesis. To determine if the outer capsid protein {sigma}3, which interacts with {micro}1, also functions in regulating cell death, we used siRNA-mediated knockdown. Similar to that observed by diminishment of {micro}1 expression, knockdown of newly synthesized {sigma}3 enhances necroptosis. {sigma}3 knockdown does not impact reovirus RNA synthesis. Instead, this increase in necroptosis following {sigma}3 knockdown is accompanied by an increase in IFN production. Furthermore, ectopic expression of {sigma}3 is sufficient to block IFN expression following infection. Surprisingly, the capacity of {sigma}3 protein to bind dsRNA does not impact its capacity to diminish production of IFN. Consistent with this, infection with a virus harboring a mutation in the dsRNA binding domain of {sigma}3 does not result in enhanced production of IFN or cell death. Together, these data suggest that {sigma}3 limits the production of IFN to control innate immune signaling and cell death following infection through a mechanism that is independent of its dsRNA binding capacity. IMPORTANCEWe use mammalian reovirus as a model to study how virus infection modulates innate immune signaling and cell death induction. Here we sought to determine how viral factors regulate these processes. Our work highlights a previously unknown role for the reovirus outer capsid protein {sigma}3 in limiting the induction of a necrotic form of cell death called necroptosis. Induction of cell death by necroptosis requires production of interferon. {sigma}3 limits the induction of necroptosis by preventing excessive production of interferon following infection.

microbiology↗

bioRxiv2020

TIRAP drives myelosuppression through an Ifnγ-Hmgb1 axis that disrupts the marrow microenvironment

Activation of inflammatory pathways is associated with bone marrow failure syndromes, but how specific molecules impact on the marrow microenvironment is not well elucidated. We report a novel role for the miR-145 target, Toll/Interleukin-1 receptor domain containing adaptor protein (TIRAP), in driving bone marrow failure. We show that TIRAP is overexpressed in various types of myelodysplastic syndromes (MDS), and suppresses all three major hematopoietic lineages.. Constitutive expression of TIRAP in hematopoietic stem/progenitor cells (HSPC) promotes upregulation of Ifn{gamma}, leading to bone marrow failure. Myelopoiesis is suppressed through Ifn{gamma}-Ifn{gamma}r-mediated release of the alarmin, Hmgb1, which disrupts the marrow endothelial niche. Deletion of Ifn{gamma} or Ifn{gamma}r blocks Hmgb1 release and is sufficient to reverse the endothelial defect and prevent myelosuppression. In contrast, megakaryocyte and erythroid production is repressed independently of the Ifn{gamma} receptor. Contrary to current dogma, TIRAP-activated Ifn{gamma}-driven marrow suppression is independent of T cell function or pyroptosis. In the absence of Ifn{gamma}, TIRAP drives myeloproliferation, implicating Ifn{gamma} in suppressing the transformation of bone marrow failure syndromes to myeloid malignancy. These findings reveal novel, non-canonical roles of TIRAP, Hmgb1 and Ifn{gamma} function in the marrow microenvironment,and provide insight into the pathophysiology of preleukemic syndromes. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=165 SRC="FIGDIR/small/967018v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@d033fborg.highwire.dtl.DTLVardef@104bab9org.highwire.dtl.DTLVardef@17ed484org.highwire.dtl.DTLVardef@1b228ed_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract: Model of proposed mechanism of TIRAP-induced BMFConstitutive TIRAP expression in marrow cells releases Ifn{gamma}, which directly impacts on megakaryocyte and erythroid production, but indirectly suppresses myelopoiesis through the release of the alarmin, Hmgb1, which disrupts the marrow endothelial compartment. C_FIG

cancer biology↗

bioRxiv2020

Mutations in TAC1B: a novel genetic determinant of clinical fluconazole resistance in C. auris

Candida auris has emerged as a multidrug-resistant pathogen of great clinical concern. Approximately 90% of clinical C. auris isolates are resistant to fluconazole, the most commonly prescribed antifungal agent, yet it remains unknown what mechanisms underpin this fluconazole resistance. To identify novel mechanisms contributing to fluconazole resistance in C. auris, the fluconazole-susceptible C. auris clinical isolate AR0387 was passaged in media supplemented with fluconazole to generate derivative strains which had acquired increased fluconazole resistance in vitro. Comparative analysis of comprehensive sterol profiles, [3H]-fluconazole uptake, sequencing of C. auris genes homologous to genes known to contribute to fluconazole resistance in other species of Candida, and the relative expression of C. auris ERG11, CDR1, and MDR1 were performed. All fluconazole-evolved derivative strains were found to have acquired mutations in the zinc-cluster transcription factor-encoding gene, TAC1B, and a corresponding increase in CDR1 expression relative to the parental clinical isolate, AR0387. Mutations in TAC1B were also identified in a set of 304 globally distributed C. auris clinical isolates representing each of the four major clades. Introduction of the most common mutation found among fluconazole-resistant clinical isolates of C. auris into the fluconazole-susceptible isolate AR0387, was confirmed to increase fluconazole resistance by 8-fold, and the correction of the same mutation in a fluconazole-resistant isolate, AR0390, decreased fluconazole MIC by 16-fold. Taken together, these data demonstrate that C. auris can rapidly acquire resistance to fluconazole in-vitro, and that mutations in TAC1B significantly contribute to clinical fluconazole resistance. IMPORTANCECandida auris is an emerging multidrug-resistant pathogen of global concern, known to be responsible for outbreaks on six continents and commonly resistant to antifungals. While the vast majority of clinical C. auris isolates are highly resistant to fluconazole, an essential part of the available antifungal arsenal, very little is known about the mechanisms contributing to resistance. In this work, we show that mutations in the transcription factor TAC1B significantly contribute to clinical fluconazole resistance. These studies demonstrate that mutations in TAC1B can arise rapidly in vitro upon exposure to fluconazole, and that a multitude of resistance-associated TAC1B mutations are present among the majority of fluconazole-resistant C. auris isolates from a global collection and appear specific to a subset of lineages or clades. Thus, identification of this novel genetic determinant of resistance significantly adds to the understanding of clinical antifungal resistance in C. auris.

microbiology↗

bioRxiv2019

Comparative genomics for the elucidation of multidrug resistance (MDR) in Candida lusitaniae

Multidrug resistance (MDR) has emerged in hospitals due to the use of several agents administered in combination or sequentially to the same individual. We reported earlier MDR in Candida lusitaniae during therapy with amphotericin B (AmB), azoles and candins. We used here comparative genomic approaches between the initial susceptible isolate and 4 other isolates with different MDR profiles. From a total of 18 non-synonymous SNPs (NSS) in genome comparisons with the initial isolate, six could be associated with MDR. One of the SNPs occurred in a putative transcriptional activator (MRR1) resulting in a V668G substitution in isolates resistant to azoles and 5-fluorocytosine (5-FC). We demonstrated by gene editing that MMR1 acted by upregulation of MFS7 (a multidrug transporter) in the presence of the V668G substitution. MFS7 itself mediated not only azole resistance but also 5-FC resistance, which represents a novel resistance mechanism for this drug class. Three other distinct NSS occurred in FKS1 (a glucan synthase that is targeted by candins) in three candin-resistant isolates. Lastly, two other NSS in ERG3 and ERG4 (ergosterol biosynthesis) resulting in non-sense mutations were revealed in AmB-resistant isolates, one of which accumulated the 2 ERG NSS. AmB-resistant isolates lacked ergosterol and exhibited sterol profiles consistent with ERG3 and ERG4 defects. In conclusion, this genome analysis combined with genetics and metabolomics helped to decipher the resistance profiles identified in this clinical case. MDR isolates accumulated 6 different mutations conferring resistance to all antifungal agents used in medicine. This case study illustrates the capacity of C. lusitaniae to rapidly adapt under drug pressure within the host.\n\nImportanceAntifungal resistance is an inevitable phenomenon when fungal pathogens get exposed to antifungal drugs. These drugs can be grouped in 4 distinct classes (azoles, candins, polyenes, pyrimidine analogs) and are used in different clinical settings. Failures in therapy implicates the sequential or combined use of these different drug classes, which can result in some cases in the development of multidrug resistance (MDR). MDR is particularly challenging in the clinic since it drastically reduces possible treatment alternatives. In this study, we report the rapid development of MDR in Candida lusitaniae in a patient, which became resistant to all known antifungal agents used up to now in medicine. To understand how MDR developed in C. lusitaniae, whole genome sequencing followed by comparative genome analysis was undertaken in sequential MDR isolates. This helped to detect all specific mutations linked to drug resistance and explained the different MDR patterns exhibited by the clinical isolates.

microbiology↗

bioRxiv2019

A framework for predicting soft-fruit yields and phenology using embedded, networked microsensors, coupled weather models and machine-learning techniques

Predicting harvest timing is a key challenge to sustainably develop soft fruit farming and reduce food waste. Soft fruits are perishable, high-value and seasonal, and sales prices are typically time-sensitive. In addition, fruit harvesting is labour-intensive and increasingly expensive making accurate phenological predictions valuable for growers. A novel approach for predicting soft fruit phenology and yields was developed and tested, using strawberries as the model crop. Seedlings were planted in polytunnels, and environmental and yield data were collected throughout the growing season. Over 1.2 million datapoints were collected by networked microsensors which measured spatial and temporal variability in air temperature, relative humidity (RH), soil moisture and photosynthetically active radiation (PAR). Fleeces were added to a subset of the plants to generate additional within-polytunnel variation. Cumulative fruit yields followed logistic growth curves and the coefficients of these curves were dependent on micro-climatic growing conditions. After 10,000 iterations, machine learning revealed that RH was the optimal factor informing the coefficients of these curves, perhaps because it is an integrative metric of air temperature and water status. Trigonometric models transformed weather forecasts, which showed a relatively low agreement with polytunnel air temperature (R2 = 0.6) and RH (R2 = 0.5) measurements, into more accurate polytunnel-specific predictions for temperature and RH (both R2 = 0.8). We present a framework for using machine-learning techniques to calculate curve coefficients and parametrise coupled weather models which can predict fruit yields and timing to a greater degree of accuracy that previously possible. Dataloggers measuring environmental and yield data could infer model parameters using iterative training for novel fruit varieties or crop types growing in different locations without a-priori phenological information. At this stage in the development of artificial intelligence and networked microsensors, this is a step forward in generating bespoke phenological prediction models to inform and support growers.

plant biology↗

bioRxiv2018

Lightweight bioinformatics: evaluating the utility of Single Board Computer (SBC) clusters for portable, scalable Real-Time Bioinformatics in fieldwork environments via benchmarking.

The versatility of the current DNA sequencing platforms and the development of portable, nanopore sequencers means that it has never been easier to collect genetic data for unknown sample ID. In fact, the distinction between fieldwork and the laboratory is becoming blurred since genome-scale data can now be collected in challenging conditions in a matter of hours. However, the full scientific and societal benefits of these new methods can only be realised with equally rapid and portable analyses. At present, field-based analyses of genomic data, despite advances in computing technology, remain problematic; laptop computers are relatively expensive and limited in scalability, while cloud- and cluster-based analyses depend, for the time being, on sufficiently reliable high-bandwidth data uplinks to transmit primary data for analysis.\n\nSingle board computers (SBCs), such as the Raspberry Pi, offer a potential solution to this problem: while less powerful than their laptop cousins, their very individual low cost and power consumption mean modest arrays of SBCs could be used for field-based preprocessing, or complete analyses or primary data. In this study we investigate the performance of one SBC, the Pi 3 Model B+, on a range of typical field-sequencing tasks versus laptop and cloud-based form-factors. Our data analysis pipeline has been made available as a workflow on Github for simple, scalable deployment for a range of uses.

bioinformatics↗

bioRxiv2018

Rapid, raw-read reference and identification (R4IDs): A flexible platform for rapid generic species ID using long-read sequencing technology.

The versatility of the current DNA sequencing platforms and the development of portable, nanopore sequencers means that it has never been easier to collect genetic data for unknown sample ID. DNA barcoding and meta-barcoding have become increasingly popular and barcode databases continue to grow at an impressive rate. However, the number of canonical genome assemblies (reference or draft) that are publically available is relatively tiny, hindering the more widespread use of genome scale DNA sequencing technology for accurate species identification and discovery. Here, we show that rapid raw-read reference datasets, or R4IDs for short, generated in a matter of hours on the Oxford Nanopore MinION, can bridge this gap and accelerate the generation of useable reference sequence data. By exploiting the long read length of this technology, shotgun genomic sequencing of a small portion of an organisms genome can act as a suitable reference database despite the low sequencing coverage. These R4IDs can then be used for accurate species identification with minimal amounts of re-sequencing effort (1000s of reads). We demonstrated the capabilities of this approach with six vascular plant species for which we created R4IDs in the laboratory and then re-sequenced, live at the Kew Science Festival 2016. We further validated our method using simulations to determine the broader applicability of the approach. Our data analysis pipeline has been made available as a Dockerised workflow for simple, scalable deployment for a range of uses.

bioinformatics↗

bioRxiv2017

Hox-logic of preadaptations for social insect symbiosis in rove beetles

How symbiotic lifestyles evolve from free-living ecologies is poorly understood. In Metazoas largest family, Staphylinidae (rove beetles), numerous lineages have evolved obligate behavioral symbioses with ants or termites. Widespread convergence of this lifestyle is thought to stem from a novel, chemically defended body plan that enables free-living species to infiltrate colonies and undergo extreme evolutionary specialization. Here we show how this innovative body plan evolved, via new Hox functions in staphylinids that remodeled the coleopteran groundplan. Using a model staphylinid, Dalotia coriaria, we reveal the Hox basis for changes in thoracic appendage morphology that shortened the beetle elytron and left the abdomen physically unprotected, selecting for an abdominal defense gland that was likely key to unlocking ant and termite societies. We present evidence that the gland evolved through a novel, combinatorial role of the abdominal Hox proteins AbdA and AbdB. These proteins function together to specify distinct gland cell types in neighboring segmental compartments, each cell type synthesizing a different class of compound--irritant, ester and solvent. Only when secreted together do these compounds constitute a bioactive secretion, providing an example of emergent chemical functionality that arises from synergy between individual gland cell types. Hox-controlled specification of glandular diversity implies a modularity in compound biosynthesis that likely catalyzed the evolvability of rove beetle chemistry, including the capacity of symbiotic taxa to produce potent compounds for host deception. This study reveals how Hox-controlled body axis modifications predispose a major animal to convergently evolve into symbionts.

evolutionary biology↗

bioRxiv2017

Field-based species identification in eukaryotes using single molecule, real-time sequencing.

Advances in DNA sequencing and informatics have revolutionised biology over the past four decades, but technological limitations have left many applications unexplored1,2. Recently, portable, real-time, nanopore sequencing (RTnS) has become available. This offers opportunities to rapidly collect and analyse genomic data anywhere3-5. However, the generation of datasets from large, complex genomes has been constrained to laboratories6,7. The portability and long DNA sequences of RTnS offer great potential for field-based species identification, but the feasibility and accuracy of these technologies for this purpose have not been assessed. Here, we show that a field-based RTnS analysis of closely-related plant species (Arabidopsis spp.)8 has many advantages over laboratory-based high-throughput sequencing (HTS) methods for species level identification-by-sequencing and de novo phylogenomics. Samples were collected and sequenced in a single day by RTnS using a portable, \"al fresco\" laboratory. Our analyses demonstrate that correctly identifying unknown reads from matches to a reference database with RTnS reads enables rapid and confident species identification. Individually annotated RTnS reads can be used to infer the evolutionary relationships of A. thaliana. Furthermore, hybrid genome assembly with RTnS and HTS reads substantially improved upon a genome assembled from HTS reads alone. Field-based RTnS makes real-time, rapid specimen identification and genome wide analyses possible. These technological advances are set to revolutionise research in the biological sciences9 and have broad implications for conservation, taxonomy, border agencies and citizen science.

evolutionary biology↗