Biology subjects

Wright, G.

Publications and source records attributed to Wright, G..

28 records · Page 2Linked to original sources

bioRxiv2020

Pollen sterols are associated with phylogenetics and environment but not with pollinators

O_LIPhytosterols are primary plant metabolites that have fundamental structural and regulatory functions. They are also essential nutrients for phytophagous insects, including pollinators, that cannot synthesize sterols. Despite the well-described composition and diversity in vegetative plant tissues, few studies have examined phytosterol diversity in pollen. C_LIO_LIWe quantified 25 pollen phytosterols in 122 plant species (105 genera, 51 families) to determine their composition and diversity across plant taxa. We searched literature and databases for plant phylogeny, environmental conditions, and pollinator guilds of the species to examine the relationships with pollen sterols. C_LIO_LI24-methylenecholesterol, sitosterol and isofucosterol were the most common and abundant pollen sterols. We found phylogenetic clustering of twelve individual sterols, total sterol content and sterol diversity, and of sterol groupings that reflect their underlying biosynthesis pathway (24 carbon alkylation, ring B desaturation). Plants originating in tropical-like climates (higher mean annual temperature, lower temperature seasonality, higher precipitation in wettest quarter) were more likely to record higher pollen sterol content. However, pollen sterol composition and content showed no clear relationship with pollinator guilds. C_LIO_LIOur study is the first to show that pollen sterol diversity is phylogenetically clustered and that pollen sterol content may adapt to environmental conditions. C_LI

biochemistry↗

bioRxiv2020

A novel human fetal liver-derived model reveals that MLL-AF4 drives a distinct fetal gene expression program in infant ALL

Although 90% of children with acute lymphoblastic leukemia (ALL) are now cured1, the prognosis of infant-ALL (diagnosis within the first year of life) remains dismal2. Infant-ALL is usually caused by a single genetic hit that arises in utero: rearrangement of the MLL/KMT2A gene (MLL-r). This is sufficient to give rise to a uniquely aggressive and treatment-refractory leukemia compared to older children with the same MLL-r3-5. The reasons for disparate outcomes in patients of different ages with identical driver mutations are unknown. This paper addresses the hypothesis that fetal-specific gene expression programs co-operate with MLL-AF4 to initiate and maintain infant-ALL. Using direct comparison of fetal and adult HSC and progenitor transcriptomes we identify fetal-specific gene expression programs in primary human cells. We show that MLL-AF4-driven infant-ALL, but not MLL-AF4 childhood-ALL, displays expression of fetal-specific genes. In a direct test of this observation, we find that CRISPR-Cas9 gene editing of primary human fetal liver cells to produce a t(4;11)/MLL-AF4 translocation replicates the clinical features of infant-ALL and drives infant-ALL-specific and fetal-specific gene expression programs. These data strongly support the hypothesis that fetal-specific gene expression programs co-operate with MLL-AF4 to initiate and maintain the distinct biology of infant-ALL.

cancer biology↗

bioRxiv2020

Structural basis for distinct inflammasome complex assembly by human NLRP1 and CARD8

Nod-like receptor (NLR) proteins activate pyroptotic cell death and IL-1 driven inflammation by assembling and activating the inflammasome complex. Closely related NLR proteins, NLRP1 and CARD8 undergo unique auto-proteolysis-dependent activation and are implicated in auto-inflammatory diseases; however, the molecular mechanisms of activation are not understood. Here we report the structural basis of how the activating domains (FIINDUPA-CARD) of NLRP1 and CARD8 self-oligomerize to trigger the assembly of distinct inflammasome complexes. Recombinant FIINDUPA-CARD of NLRP1 forms a two-layered filament, with an inner core composed of oligomerized CARD domains and the outer layer consisting of FIINDUPA rings. Biochemically, oligomerized NLRP1-CARD is sufficient to drive ASC speck formation in cultured human cells via filament formation-a process that is greatly enhanced by NLRP1-FIINDUPA, which forms ring-like oligomers in vitro. In addition, we report the cryo-EM structures of NLRP1-CARD and CARD8-CARD filaments at 3.7 [A], which uncovers unique structural features that enable NLRP1 and CARD8 to discriminate between ASC and pro-caspase-1. In summary, our findings provide unique structural insight into the mechanisms of activation for human NLRP1 and CARD8, uncovering an unexpected level of specificity in inflammasome signaling mediated by heterotypic CARD domain interactions.

immunology↗

bioRxiv2020

The extent and impact of variation in ADME genes in sub-Saharan African populations

Investigating variation in genes involved in the absorption, distribution, metabolism, and excretion (ADME) of drugs are key to characterising pharmacogenomic (PGx) relationships. ADME gene variation is relatively well characterised in European and Asian populations, but African populations are under-studied - which has implications for safe and effective drug use in Africa. The genetic diversity of ADME genes across sub-Saharan African populations is large. The Southern African population cluster is most distinct from that of far West Africa. PGx strategies based on European variants will be of limited use in African populations. Although established variants are important, PGx must take into account the full range of African variation. This work urges further characterisation of variants in African populations including in vitro and in silico studies, and to consider the unique African ADME landscape when developing precision medicine guidelines and tools for African populations. Author summaryThe ADME genes are a group of genes that play a key role in absorption, distribution, metabolism and excretion of drugs. Variations in these genes can have a significant impact on drug safety and efficacy. Africa has a high level of genetic variation and is under-studied in drug development, which makes study of variations in these genes in African populations very important. Using a new data set of 458 high-coverage genomes from across Africa, we characterise the extent and impact of variation in the ADME genes, looking at both single nucleotide and copy number variations. We identified 343,368 variants, including 40,692 novel variants, and 930 coding variants which are predicted to have high impact on function. Our discovery curves indicate that there will be considerable value in sequencing more African genomes. Moreover, relatively few of these novel variants are captured on common genotyping arrays. We show that there is considerable diversity within Africa in some important genes, and this will have significant consequences for the emerging field of precision medicine in Africa.

genetics↗

bioRxiv2020

Analysis of computational codon usage models and their association with translationally slow codons

Improved computational modeling of protein translation rates, including better prediction of where translational slowdowns along an mRNA sequence may occur, is critical for understanding co-translational folding. Because codons within a synonymous codon group are translated at different rates, many computational translation models rely on analyzing synonymous codons. Some models rely on genome-wide codon usage bias (CUB), believing that globally rare and common codons are the most informative of slow and fast translation, respectively. Others use the CUB observed only in highly expressed genes, which should be under selective pressure to be translated efficiently (and whose CUB may therefore be more indicative of translation rates). No prior work has analyzed these models for their ability to predict translational slowdowns. Here, we evaluate five models for their association with slowly translated positions as denoted by two independent ribosome footprint (RFP) count experiments from S. cerevisiae, because RFP data is often considered as a "ground truth" for translation rates across mRNA sequences. We show that all five considered models strongly associate with the RFP data and therefore have potential for estimating translational slowdowns. However, we also show that there is a weak correlation between RFP counts for the same genes originating from independent experiments, even when their experimental conditions are similar. This raises concerns about the efficacy of using current RFP experimental data for estimating translation rates and highlights a potential advantage of using computational models to understand translation rates instead.

bioinformatics↗

bioRxiv2020

Developmental differentiation in a cytoplasm-dwelling obligate intracellular bacterium

Orientia tsutsugamushi (Ot) is an obligate intracellular bacterium in the family Rickettsiaceae that causes scrub typhus, a severe mite-borne human disease. Its mechanism of cell exit is unusual amongst Rickettsiaceae, as Ot buds off the surface of infected cells enveloped in plasma membrane. Here, we show that Ot bacteria that have budded out of host cells are in a distinct developmental stage compared with intracellular bacteria. We refer to these two stages as intracellular and extracellular bacteria (IB and EB, respectively). These two forms differ in physical properties: IB is elongated, and EB is round. Additionally, IB has higher levels of peptidoglycan and is physically robust compared with EB. The two bacterial forms differentially express proteins involved in bacterial physiology and host-pathogen interactions, specifically those involved in bacterial dormancy and stress response, secreted bacterial effectors, and outer membrane autotransporter proteins ScaA and ScaC. Whilst both populations are infectious, entry of IB Ot is sensitive to inhibitors of both clathrin-mediated endocytosis and macropinocytosis, whereas entry of EB Ot is only sensitive to a macropinocytosis inhibitor. Our identification and detailed characterization of two developmental forms of Ot significantly advances our understanding of the intracellular lifecycle of an important human pathogen. Author SummaryOrientia tsutsugamushi (Ot) is a bacterial pathogen that causes scrub typhus, a mite-transmitted human disease. This illness is traditionally known to be endemic in the Asia-Pacific, but recent reports of Orientia-like organisms from the Middle East, Africa, and Latin America suggest that it may be globally distributed. Scrub typhus is associated with high mortality if not treated promptly with appropriate antibiotics. Ot is a highly specialized bacterium that can only replicate within living cells, either within the mite vector or in mammalian or human hosts. Ot exits infected cells using a unique mechanism that involves budding off the surface of infected cells. We have discovered that this unusual aspect of its lifecycle involves the bacteria themselves differentiating into a distinct growth form. Different growth forms have not been described in other members of the family Rickettsiaceae, and no other family members have been shown to bud out of host cells in a manner similar to Ot. We find that the two forms of Ot, which we refer to as intracellular and extracellular bacteria (IB and EB respectively), differ in physical properties and protein expression and infect cells through different mechanisms. The identification of structurally and functionally distinct forms of Ot elucidates a vital aspect of this pathogens intracellular life cycle. The two forms are likely to have different antibiotic susceptibilities, therefore our findings may advance the development of novel interventions aimed at inhibiting Ot growth in scrub typhus patients.

microbiology↗

bioRxiv2020

Single-molecule fluorescence-based approach reveals novel mechanistic insights into small heat shock protein chaperone function

Small heat shock proteins (sHsps) are a family of ubiquitous intracellular molecular chaperones that are up-regulated under stress conditions and play a vital role in protein homeostasis (proteostasis). It is commonly accepted that these chaperones work by trapping misfolded proteins to prevent their aggregation, however fundamental questions regarding the molecular mechanism by which sHsps interact with misfolded proteins remain unanswered. Traditionally, it has been difficult to study sHsp function due to the dynamic and heterogenous nature of the species formed between sHsps and aggregation-prone proteins. Single-molecule techniques have emerged as a powerful tool to study dynamic protein complexes and we have therefore developed a novel single-molecule fluorescence-based approach to observe the chaperone action of human B-crystallin (Bc, HSPB5). Using this approach we have, for the first time, determined the stoichiometries of complexes formed between Bc and a model client protein, chloride intracellular channel 1 (CLIC1). By examining the polydispersity and stoichiometries of these complexes over time, and in response to different concentrations of Bc, we have uncovered unique and important insights into a two-step mechanism by which Bc interacts with misfolded client proteins to prevent their aggregation. Understanding this fundamental mechanism of sHsp action is crucial to understanding how these molecular chaperone function to protect the cell from protein misfolding and their overall role in the cellular proteostasis network.

biochemistry↗

bioRxiv2020

Mechanical Instability of Adherens Junctions Overrides Intrinsic Quiescence of Hair Follicle Stem Cells

Vinculin, a mechanotransducer associated with both adherens junctions (AJ) and focal adhesions (FA) plays a central role in force transmission through these cell-cell and cell-substratum contacts. Here we describe the conditional knock out (KO) of vinculin in murine skin. Remarkably, we find that the loss of vinculin function results in the loss of bulge stem cell (BuSC) quiescence. We demonstrate that vinculin KO cells are impaired in force generation resulting in mechanically weak AJs. Mechanistically, vinculin functions by keeping -catenin in a stretched conformation, which in turn regulates the retention of YAP1, another potent mechanotransducer and regulator of cell proliferation, to the junctions. Conditional KO of -catenin specifically in the BuSCs further corroborates the importance of stable AJs in the maintenance of quiescence and stemness. Altogether, our data provides definitive mechanistic insights into the hitherto unexplored regulatory link between the mechanical stability of cell-junctions and the maintenance of BuSC quiescence.

cell biology↗

bioRxiv2018

Genome location dictates the transcriptional response to sub-inhibitory concentrations of PolC-inhibitors in Clostridium difficile

Clostridium difficile is a potentially lethal gut pathogen that causes nosocomial and community acquired infections. Limited treatment options and reports of reduced susceptibility to current treatment emphasize the necessity for novel antimicrobials. The DNA-polymerase of gram-positive organisms is an attractive target for the development of antimicrobials. ACX-362E (N2-(3<,4-Dichlorobenzyl)-7-(2-[1-morpholinyl]ethyl)guanine; MorE-DCBG) is a DNA polymerase inhibitor in pre-clinical development as a novel therapeutic against C. difficile infection. This synthetic purine shows preferential activity against C. difficile PolC over those of other organisms in vitro and is effective in an animal model of C. difficile infection. In this study we have determined its efficacy against a large collection of clinical isolates. At concentrations below the minimal inhibitory concentration, the presumed slowing (or stalling) of replication forks due to ACX-362E leads to a growth defect. We have determined the transcriptional response of C. difficile to replication inhibition and observed an overrepresentation of up-regulated genes near the origin of replication in the presence of PolC-inhibitors, but not when cells were subjected to sub-inhibitory concentrations of other antibiotics. This phenomenon can be explained by a gene dosage shift, as we observed a concomitant increase in the ratio between origin-proximal versus terminus-proximal gene copy number upon exposure to PolC-inhibitors. Moreover, we show that certain genes differentially regulated under PolC-inhibition are controlled by the origin-proximal general stress response regulator sigma factor B. Together, these data suggest that genome location both directly and indirectly determines the transcriptional response to replication inhibition in C. difficile.

microbiology↗

bioRxiv2017

Mesophotic Coral Ecosystems Inside and Outside a Caribbean Marine Protected Area

Recent widespread shallow coral reef loss has led to calls for more holistic approaches to coral reef management, requiring inclusion of all ecosystems interacting with coral reefs in management plans. Yet almost all current reef management is biased towards shallow reefs, and overlooks that many reef species can also be found on mesophotic coral ecosystems (MCEs; reefs 30 -150 m). This study presents the first detailed quantitative characterisation of MCEs off Cozumel, in the Mexican Caribbean and provides insights into their general state. We investigate whether MCEs within the marine park have similar ecological communities to mesophotic reefs outside protection, despite widely recognised shallow reef impacts outside the protected area. Results show some taxon specific differences in MCE benthic communities between sites within the protected area and areas outside; although overall communities are similar. Regardless of protection and location, and in contrast to shallow reefs, all observed Cozumel MCEs were continuous reefs dominated by calcareous macroalgae, sponges, octocorals, and black corals. Hard corals were present on MCEs, but at low abundance. We found that 42.5 % of fish species recorded on Cozumel could be found on both shallow reefs and MCEs, including many commercially-important fish species. This suggest that MCEs may play a role in supporting fish populations. However, regardless of protection status and depth we found that large-body fishes (>500 mm) were nearly absent at all studied sites. MCEs should be incorporated into the existing shallow-reef focused management plan in Cozumel, with well informed and implemented fisheries and harvesting regulations.

ecology↗