Biology subjects

Jones, A.

Publications and source records attributed to Jones, A..

At least 91 records · Page 5Linked to original sources

bioRxiv2022

An assessment of bioinformatics tools for the detection of human endogenous retroviral insertions in short-read genome sequencing data

There is a growing interest in the study of human endogenous retroviruses (HERVs) given the substantial body of evidence that implicates them in many human diseases. Although their genomic characterization presents numerous technical challenges, next-generation sequencing (NGS) has shown potential to detect HERV insertions and their polymorphisms in humans, and a number of computational tools to detect them in short-read NGS data exist. In order to design optimal analysis pipelines, an independent evaluation of the currently available tools is required. We evaluated the performance of a set of such tools using a variety of experimental designs and types of NGS datasets. These included 50 human short read whole-genome sequencing samples, matching long and short read NGS data, and simulated short-read NGS data. Our results highlight the performance variability of the tools across the datasets and suggest that different tools might be suitable for different study designs. Using multiple tools and a consensus approach is advisable if computationally feasible and wet-lab validation via PCR is advisable where biological samples are available.

bioinformatics↗

bioRxiv2022

Alignment of spatial genomics and histology data using deep Gaussian processes

Spatially-resolved genomic technologies have allowed us to study the physical organization of cells and tissues, and promise an understanding of the local interactions between cells. However, it remains difficult to precisely align spatial observations across slices, samples, scales, individuals, and technologies. Here, we propose a probabilistic model that aligns a set of spatially-resolved genomics and histology slices onto a known or unknown common coordinate system into which the samples are aligned both spatially and in terms of the phenotypic readouts (e.g., gene or protein expression levels, cell density, open chromatin regions). Our method consists of a two-layer Gaussian process: the first layer maps the observed samples spatial locations into a common coordinate system, and the second layer maps from the common coordinate system to the observed readouts. Our approach also allows for slices to be mapped to a known template coordinate space if one exists. We show that our registration approach enables complex downstream spatially-aware analyses of spatial genomics data at multiple resolutions that are impossible or inaccurate with unaligned data, including an analysis of variance, differential expression across the z-axis, and association tests across multiple data modalities.

bioinformatics↗

bioRxiv2021

Cellular heterogeneity of human fallopian tubes in normal and hydrosalpinx disease states identified by scRNA-seq

Fallopian tube (FT) homeostasis requires dynamic regulation of heterogeneous cell populations and is disrupted in infertility and ovarian cancer. Here we applied single-cell RNAseq to profile 53,376 FT cells from 3 healthy pre-menopausal subjects. The resulting cell atlas contains 12 major cell types representing epithelial, stromal and immune compartments. Re-clustering of epithelial cells identified 4 ciliated and 6 non-ciliated secretory epithelial subtypes, two of which represent potential progenitor pools: one leading to mature secretory cells, while the other contributing to either ciliated cells or one of the stromal cell types. To understand how FT cell numbers and states change in a disease state, we analyzed ~15,000 cells from a hydrosalpinx sample and observed shifts in epithelial and stromal populations, and cell type-specific changes in extracellular matrix and TGF-{beta} signaling, underscoring fibrosis pathophysiology. This resource is expected to facilitate future studies to understand fallopian tube homeostasis in normal development and disease.

systems biology↗

bioRxiv2021

Genomic constraints to drought adaptation

Global shifts in climate and precipitation patterns are altering the diversity and structure of forests. The ability for species to adapt is especially difficult for long lived foundation species with unknown genetic and trait diversity. We harnessed genomic, physiological, and climate data to determine adaptation constraints. We used denovo assembly and 6.5 million genomic variants with drought related traits from 432 individuals sourced from across the complete range of the foundation tree species Corymbia calophylla. We found genomic variants determining traits predominantly in gene regulatory regions. The ability for populations to adapt was limited by within population genetic diversity associated with traits, and epistatic interactions within traits and pleiotropic interactions among traits. Nevertheless, we could accurately predict adaptive traits using genomic and climate data to guide reforestation. Our study indicated that some populations may contain variation sufficient for the species to adapt to the effects of drought, while other populations will need increased variability from those sources.

evolutionary biology↗

bioRxiv2021

A multi-gene region targeted capture approach to detect plant DNA in environmental samples: A case study from coastal environments

Metabarcoding of plant DNA recovered from environmental samples, termed environmental DNA (eDNA), has been used to detect invasive species, track biodiversity changes and reconstruct past ecosystems. The P6 loop of the trnL intron is the most widely utilized gene region for metabarcoding plants due to the short fragment length and subsequent ease of recovery from degraded DNA, which is characteristic of environmental samples. However, the taxonomic resolution for this gene region is limited, often precluding species level identification. Additionally, targeting gene regions using universal primers can bias results as some taxa will amplify more effectively than others. To increase the ability of DNA metabarcoding to better resolve flowering plant species (angiosperms) within environmental samples, and reduce bias in amplification, we developed a multi-gene targeted capture method that simultaneously targets 20 chloroplast gene regions in a single assay across all flowering plant species. Using this approach, we effectively recovered multiple chloroplast gene regions for three species within artificial DNA mixtures down to 0.001 ng/{micro}L of DNA. We tested the detection level of this approach, successfully recovering target genes for 10 flowering plant species. Finally, we applied this approach to sediment samples containing unknown compositions of environmental DNA and confidently detected plant species that were later verified with observation data. Targeting multiple chloroplast gene regions in environmental samples enabled species-level information to be recovered from complex DNA mixtures. Thus, the method developed here, confers an improved level of data on community composition, which can be used to better understand flowering plant assemblages in environmental samples.

molecular biology↗

bioRxiv2021

FoxA1 and FoxA2 regulate growth and cellular identity in NKX2-1-positive lung adenocarcinoma

Changes in cellular identity (also known as histologic transformation or lineage plasticity) can drive malignant progression and resistance to therapy in multiple types of cancer, including lung adenocarcinoma (LUAD). The lineage specifying transcription factors FoxA1 and FoxA2 (FoxA1/2) regulate identity in NKX2-1/TTF1-negative LUAD. However, their role in NKX2-1-positive LUAD has not been systematically investigated. We find that Foxa1/2 knockout severely impairs tumorigenesis in KRAS-driven genetically engineered mouse models and human cell lines. Foxa1/2 deletion leads to the collapse of a dual identity state, marked by co-expression of pulmonary and gastrointestinal transcriptional programs, which has been implicated in LUAD progression. Mechanistically, loss of FoxA1/2 leads to aberrant NKX2-1 activity and genomic localization, which inhibits tumorigenesis and drives alternative cellular identity programs associated with non-proliferative states. This work demonstrates that FoxA1/2 expression is a novel lineage-specific vulnerability in NKX2-1-positive LUAD and identifies mechanisms of response and resistance to targeting FoxA1/2 in this disease.

cancer biology↗

bioRxiv2021

The K18-hACE2 Transgenic Mouse Model Recapitulates Non-Severe and Severe COVID-19 in Response to Infectious Dose of SARS-CoV-2 Virus

A comprehensive analysis and characterization of a SARS-CoV-2 infection model that mimics non-severe and severe COVID-19 in humans is warranted for understating the virus and developing preventive and therapeutic agents. Here, we characterized the K18-hACE2 mouse model expressing human (h)ACE2 in mice, controlled by the human keratin 18 (K18) promoter, in epithelia, including airway epithelial cells where SARS-CoV-2 infections typically start. We found that intranasal inoculation with higher viral doses (2x103 and 2x104 PFU) of SARS-CoV-2 caused lethality of all mice and severe damage of various organs, including lungs, liver, and kidney, while lower doses (2x101 and 2x102 PFU) led to less severe tissue damage and some mice recovered from the infection. In this humanized hACE2 mouse model, SARS-CoV-2 infection damaged multiple tissues, with a dose-dependent effect in most tissues. Similar damage was observed in biopsy samples from COVID-19 patients. Finally, the mice that recovered after infection with a low dose of virus also survived rechallenge with a high dose of virus. Compared to other existing models, the K18-hACE2 model seems to be the most sensitive COVID-19 model reported to date. Our work expands the information available about this model to include analysis of multiple infectious doses and various tissues with comparison to human biopsy samples from COVID-19 patients. In conclusion, the K18-hACE2 mouse model recapitulates both severe and non-severe COVID-19 in humans and can provide insight into disease progression and the efficacy of therapeutics for preventing or treating COVID-19. ImportanceThe pandemic of COVID-19 has reached 112,589,814 cases and caused 2,493,795 deaths worldwide as of February 23, 2021, has raised an urgent need for development of novel drugs and therapeutics to prevent the spread and pathogenesis of SARS-CoV-2. To achieve this goal, an animal model that recapitulates the features of human COVID-19 disease progress and pathogenesis is greatly needed. In this study, we have comprehensively characterized a mouse model of SARS-CoV-2 infection using K18-hACE2 transgenic mice. We infected the mice with low and high doses of SARS-CoV-2 virus to study the pathogenesis and survival in response to different infection patterns. Moreover, we compared the pathogenesis of the K18-hACE2 transgenic mice with that of the COVID-19 patients to show that this model could be a useful tool for the development of anti-viral drugs and therapeutics.

microbiology↗

bioRxiv2021

Identification and correction of phase switches with Hi-C data in the Nanopore and HiFi chromosome-scale assemblies of the dikaryotic leaf rust fungus Puccinia triticina

BackgroundMost animals and plants have more than one set of chromosomes and package these haplotypes into a single nucleus within each cell. In contrast, many fungal species carry multiple haploid nuclei per cell. Rust fungi are such species with two nuclei (karyons) that contain a full set of haploid chromosomes each. The physical separation of haplotypes in dikaryons means that, unlike in diploids, Hi-C chromatin contacts between haplotypes are false positive signals. ResultsWe generate the first chromosome-scale, fully-phased assembly for the dikaryotic leaf rust fungus Puccinia triticina and compare Nanopore MinION and PacBio HiFi sequence-based assemblies. We show that false positive Hi-C contacts between haplotypes are predominantly caused by phase switches rather than by collapsed regions or Hi-C read mis-mappings. We introduce a method for phasing of dikaryotic genomes into the two haplotypes using Hi-C contact graphs, including a phase switch correction step. In the HiFi assembly, relatively few phase switches occur, and these are predominantly located at haplotig boundaries and can be readily corrected. In contrast, phase switches are widespread throughout the Nanopore assembly. We show that haploid genome read coverage of 30-40 times using HiFi sequencing is required for phasing of the leaf rust genome (~0.7% heterozygosity) and that HiFi sequencing resolves genomic regions with low heterozygosity that are otherwise collapsed in the Nanopore assembly. ConclusionsThis first Hi-C based phasing pipeline for dikaryons and comparison of long-read sequencing technologies will inform future genome assembly and haplotype phasing projects in other non-haploid organisms.

genomics↗

bioRxiv2020

In vivo proteomic mapping through GFP-directed proximity dependent biotin labelling in zebrafish

Protein interaction networks are crucial for complex cellular processes. However, the elucidation of protein interactions occurring within highly specialised cells and tissues is challenging. Here we describe the development, and application, of a new method for proximity-dependent biotin labelling in whole zebrafish. Using a conditionally stabilised GFP-binding nanobody to target a biotin ligase to GFP-labelled proteins of interest, we show tissue-specific proteomic profiling using existing GFP-tagged transgenic zebrafish lines. We demonstrate the applicability of this approach, termed BLITZ (Biotin Labelling In Tagged Zebrafish), in diverse cell types such as neurons and vascular endothelial cells. We applied this methodology to identify interactors of caveolar coat protein, cavins, in skeletal muscle. Using this system, we defined specific interaction networks within in vivo muscle cells for the closely related but functionally distinct Cavin4 and Cavin1 proteins.

cell biology↗

bioRxiv2020

SETD2 regulates the methylation of translation elongation factor eEF1A1 in clear cell renal cell carcinoma

SET domain-containing protein 2 (SETD2) is commonly mutated in renal cell carcinoma. SETD2 methylates histone H3 as well as a growing list of non-histone proteins. To explore SETD2-dependent regulation of the kidney cancer proteome, we performed a systems-wide analysis of protein lysine-methylation and expression in wild type (WT) and SETD2-knock out (KO) kidney cells. We observed decreased lysine methylation of the translation elongation factor eEF1A1. EEF1AKMT2 and EEF1AKMT3 are known to methylate eEF1A1, and we show here that their expression is dependent on SET-domain function of SETD2. Globally, we observe differential expression of hundreds of proteins in WT versus SETD2-KO cells, including increased expression of many involved in protein translation. Finally, we observe decreased progression free survival and loss of EEF1AKMT2 gene expression in SETD2-mutated tumors. Overall, these data suggest that SETD2-mutated ccRCC, via loss of enzymetic function of the SET domain, displays dysregulation of protein translation as a potentially important component of the transformed phenotype.

cancer biology↗

bioRxiv2020

Targeting Vitamin-D receptor (VDR) by a small molecule antagonist MeTC7 inhibits PD-L1 but controls THMYCN neuroblastoma growth PD-L1 independently

Vitamin-D receptor (VDR) mRNA is enriched in malignant lung, ovarian and pancreatic tissues and showed poor prognoses. Calcitriol and stable or CRISPR-directed VDR upregulation increased PD-L1mRNA and protein expression in cancer cells in-vitro. A ChIP assay showed the binding of VDR with VDREPD-L1. Stattic, a STAT3 phosphorylation inhibitor blocked calcitriol or VDR overexpression induced PD-L1 upregulation. MeTC7, a VDR antagonist developed by us, reduced PD-L1 expression on macrophages, ovarian, lung, breast, and pancreatic cancer cells in-vitro. In radiotherapy inducible PD-L1 model of orthotopic MC38 murine colon cancer, MeTC7 decreased PD-L1 surface expression, suppressed inflammatory monocytes (IMs) population and increased intra-tumoral CD69+PD1+CD8+T-cells. Intriguingly, MeTC7 reduced TH-MYCN transgenic neuroblastoma tumor growth without affecting PD-L1 and tumor immune milieu. In summary, Vitamin-D/VDR drives PD-L1 expression on cancer cells via STAT-3. Inhibiting VDR exhibited anti-checkpoint effects in orthotopic colon tumors, whereas PDL1-independent and anti-VDR/MYCN effects controlled growth of transgenic neuroblastoma and xenografted tumors. SummaryVitamin-D/VDR induces PD-L1 expression on cancer cells via STAT-3; and targeting VDR by a novel small molecule antagonist MeTC7 exhibits both anti-PD-L1 and anti-VDR/MYCN effects in tumor models.

cancer biology↗

bioRxiv2020

The RNA-binding protein Igf2bp3 is critical for embryonic and germline development in zebrafish

The ability to reproduce is essential in all branches of life. In metazoans, this process is initiated by formation of the germline, a group of cells that are destined to form the future gonads, the tissue that will produce the gametes. The molecular mechanisms underlying germline formation differs between species. In zebrafish, development of the germline is dependent on the specification, migration and proliferation of progenitors called the primordial germ cells (PGCs). PGC specification is dependent on a maternally provided cytoplasmic complex of ribonucleoproteins (RNPs), the germplasm. Here, we show that the conserved RNA-binding protein (RBP), Igf2bp3, has an essential role during germline development. Loss of Igf2bp3 leads to reduced germline RNA expression, and mis-regulated germline development. Maternal mutants affecting igf2bp3 exhibit abnormal PGC movement, maintenance and migration, and adult igf2bp3 mutants show a male biased sex ratio. Therefore, Igf2bp3 is required for germplasm components, and maintains correct development of the germline.Competing Interest StatementThe authors have declared no competing interest.View Full Text

developmental biology↗

bioRxiv2020

Selective vulnerability of aneuploid human cancer cells to inhibition of the spindle assembly checkpoint

Selective targeting of aneuploid cells is an attractive strategy for cancer treatment. Here, we mapped the aneuploidy landscapes of ~1,000 human cancer cell lines and classified them by their degree of aneuploidy. Next, we performed a comprehensive analysis of large-scale genetic and chemical perturbation screens, in order to compare the cellular vulnerabilities between near-diploid and highly-aneuploid cancer cells. We identified and validated an increased sensitivity of aneuploid cancer cells to genetic perturbation of core components of the spindle assembly checkpoint (SAC), which ensures the proper segregation of chromosomes during mitosis. Surprisingly, we also found highly-aneuploid cancer cells to be less sensitive to short-term exposures to multiple inhibitors of the SAC regulator TTK. To resolve this paradox and to uncover its mechanistic basis, we established isogenic systems of near-diploid cells and their aneuploid derivatives. Using both genetic and chemical inhibition of BUB1B, MAD2 and TTK, we found that the cellular response to SAC inhibition depended on the duration of the assay, as aneuploid cancer cells became increasingly more sensitive to SAC inhibition over time. The increased ability of aneuploid cells to slip from mitotic arrest and to keep dividing in the presence of SAC inhibition was coupled to aberrant spindle geometry and dynamics. This resulted in a higher prevalence of mitotic defects, such as multipolar spindles, micronuclei formation and failed cytokinesis. Therefore, although aneuploid cancer cells can overcome SAC inhibition more readily than diploid cells, the proliferation of the resultant aberrant cells is jeopardized. At the molecular level, analysis of spindle proteins identified a specific mitotic kinesin, KIF18A, whose levels were drastically reduced in aneuploid cancer cells. Aneuploid cancer cells were particularly vulnerable to KIF18A depletion, and KIF18A overexpression restored the sensitivity of aneuploid cancer cells to SAC inhibition. In summary, we identified an increased vulnerability of aneuploid cancer cells to SAC inhibition and explored its cellular and molecular underpinnings. Our results reveal a novel synthetic lethal interaction between aneuploidy and the SAC, which may have direct therapeutic relevance for the clinical application of SAC inhibitors.

cancer biology↗

bioRxiv2020

What makes Hemidactylus invasions successful? A case study on the island of Curacao.

Hemidactylus spp. (House geckos) rank among the most successful invasive reptile species worldwide. Hemidactylus mabouia in particular has become ubiquitous across tropical urban settings in the Western Hemisphere. H. mabouia’s ability to thrive in close proximity to humans has led to the rapid displacement of native geckos in urban areas, however the mechanisms driving this displacement remain understudied. Here we combine data from nitrogen and carbon stable isotopes, stomach contents, and morphometric analyses of traits associated with feeding and locomotion to test alternate hypotheses of displacement between H. mabouia and a native gecko, Phyllodactylus martini, on the island of Curaçao. Consistent with expectations of direct food resource competition, we demonstrate substantial overlap of invertebrate prey resources between the species. Additionally, we found strong evidence from both diet content and stable isotope analyses that H. mabouia acts as a vertebrate predator, preying upon P. martini as well as other native and non-native reptiles. Finally, we show that H. mabouia possesses several morphological advantages, including larger sizes in feeding-associated traits and limb proportions that could offer a propulsive locomotor advantage on vertical surfaces. Together, these findings suggest the successful establishment of H. mabouia likely involves a combination of both exploitative interspecific competition and predation. Given the ubiquity of H. mabouia, illuminating the role of this species as both a competitor and a predator casts new concerns on the ecological and demographic impacts of this widespread urban invader.Competing Interest StatementThe authors have declared no competing interest.View Full Text

zoology↗

bioRxiv2020

Global computational alignment of tumor and cell line transcriptional profiles

Cell lines are key tools for preclinical cancer research, but it remains unclear how well they represent patient tumor samples. Identifying cell line models that best represent the features of particular tumor samples, as well as tumor types that lack in vitro model representation, remain important challenges. Gene expression has been shown to provide rich information that can be used to identify tumor subtypes, as well as predict the genetic dependencies and chemical vulnerabilities of cell lines. However, direct comparisons of tumor and cell line transcriptional profiles are complicated by systematic differences, such as the presence of immune and stromal cells in tumor samples and differences in the cancer-type composition of cell line and tumor expression datasets. To address these challenges, we developed an unsupervised alignment method (Celligner) and applied it to integrate several large-scale cell line and tumor RNA-Seq datasets. While our method aligns the majority of cell lines with tumor samples of the same cancer type, it also reveals large differences in tumor/cell line similarity across disease types. Furthermore, Celligner identifies a distinct group of several hundred cell lines from diverse lineages that present a more mesenchymal and undifferentiated transcriptional state and which exhibit distinct chemical and genetic dependencies. This method could thus be used to guide the selection of cell lines that more closely resemble patient tumors and improve the clinical translation of insights gained from cell line models.

cancer biology↗

bioRxiv2020

Long read assembly of the pandemic strain of Austropuccinia psidii (myrtle rust) reveals an unusually large (gigabase sized) and repetitive fungal genome.

Austropuccinia psidii, originating in South America, is a globally invasive fungal plant pathogen that causes rust disease on Myrtaceae. Several biotypes are recognized, with the most widely distributed pandemic biotype spreading throughout the Asia-Pacific and Oceania regions over the last decade. Austropuccinia psidii has a broad host range with more than 480 myrtaceous species. Since first detected in Australia in 2010, the pathogen has caused the near extinction of at least three species and negatively affected commercial production of several Myrtaceae. To enable molecular and evolutionary studies into A. psidii pathogenicity, we assembled a highly contiguous genome for the pandemic biotype. With an estimated haploid genome size of just over 1 Gb (gigabases), it is the largest assembled fungal genome to date. The genome has undergone massive expansion via distinct transposable element (TE) bursts. Over 90% of the genome is covered by TEs predominantly belonging to the Gypsy superfamily. These TE bursts have likely been followed by deamination events of methylated cytosines to silence the repetitive elements. This in turn led to the depletion of CpG sites in transposable elements and a very low overall GC content of 33.8%. The overall gene content is highly conserved, when compared to other closely related Pucciniales, yet the intergenic distances are increased by an order of magnitude indicating a general insertion of TEs between genes. Overall, we show how transposable elements shaped the genome evolution of A. psidii and provide a greatly needed resource for strategic approaches to combat disease spread.

genomics↗

bioRxiv2020

Post-perturbational transcriptional signatures of cancer cell line vulnerabilities

While chemical and genetic viability screens in cancer cell lines have identified many promising cancer vulnerabilities, simple univariate readouts of cell proliferation fail to capture the complex cellular responses to perturbations. Complementarily, gene expression profiling offers an information-rich measure of cell state that can provide a more detailed account of cellular responses to perturbations. Relatively little is known, however, about the relationship between transcriptional responses to per-turbations and the long-term cell viability effects of those perturbations. To address this question, we integrated thousands of post-perturbational transcriptional profiles from the Connectivity Map with large-scale screens of cancer cell lines viability response to genetic and chemical perturbations. This analysis revealed a generalized transcriptional signature associated with reduced viability across perturbations, which was consistent across post-perturbation time-points, perturbation types, and viability datasets. At a more granular level, we lay out the landscape of treatment-specific expression-viability relationships across a broad panel of drugs and genetic reagents, and we demonstrate that these post-perturbational expression signatures can be used to infer long-term viability. Together, these results help unmask the transcriptional changes that are associated with perturbation-induced viability loss in cancer cell lines.

bioinformatics↗

bioRxiv2020

An NKX2-1/ERK/WNT feedback loop modulates gastric identity and response to targeted therapy in lung adenocarcinoma

Cancer cells undergo lineage switching during natural progression and in response to therapy. NKX2-1 loss in human and murine lung adenocarcinoma leads to invasive mucinous adenocarcinoma (IMA), a lung cancer subtype that exhibits gastric differentiation and harbors a distinct spectrum of driver oncogenes. NKX2-1 is required for optimal tumor initiation, but dispensable for growth of established tumors, in BRAFV600E driven disease. NKX2-1-deficient, BRAFV600E driven tumors resemble human IMA and exhibit a distinct response to BRAF/MEK inhibitors. Whereas BRAF/MEK inhibitors drive NKX2-1-positive tumor cells into quiescence, NKX2-1-negative cells fail to exit the cell cycle after the same therapy. BRAF/MEK inhibitors also induce cell identity switching in NKX2-1-negative lung tumors within the gastric lineage, which is driven in part by WNT signaling and FoxA1/2. These data elucidate a complex, reciprocal relationship between lineage specifiers and oncogenic signaling pathways in the regulation of lung adenocarcinoma identity that is likely to impact lineage-specific therapeutic strategies.

cancer biology↗