Biology subjects

Bottrill, A.

Publications and source records attributed to Bottrill, A..

11 recordsLinked to original sources

bioRxiv2026

Plasmodium rootlet SFA1 coordinates cell replication, apical polarity and cytoskeletal organisation during sexual development

Rootlet fibres are evolutionarily conserved cytoskeletal structures involved in basal body positioning and cytoskeletal organisation in diverse eukaryotes. Plasmodium spps, undergo complex developmental transitions that require precise coordination of cell polarity, cytoskeletal remodelling and nuclear division. In Plasmodium striated fibre assemblin1 (SFA1) and SFA2 have been implicated in linking the centriolar plaque to nascent apical poles during schizogony and sporogony. However, their role in sexual development is unclear. Here we show SFA1 is absent from the male basal body but is present in female gametocytes, where it marks the polarity initiation site during zygote-to-ookinete differentiation. We also identify associations between SFA1, SFA2 and pellicle-associated proteins. Loss of sfa1 disrupts apical polarity, pellicle organisation and subpellicular microtubules, producing abnormal immotile ookinetes and affecting further sporozoite development, leading to complete loss of parasite transmission. Together, these findings support an important role for SFA1 in organising the cytoskeletal architecture required for malaria transmission.

cell biology↗

bioRxiv2026

GAPMs form a heterotrimeric complex bridging the gliding machinery and the cytoskeleton across Plasmodium species

Apicomplexan parasites, such as malaria-causing Plasmodium spp., use a specialised actomyosin motor system known as the glideosome to drive movement through host tissue and invade host cells. This system is anchored to the inner membrane complex (IMC), a series of flattened vesicles located beneath the plasma membrane, and thought to be linked to the underlying cytoskeleton by the GAPM protein family. However, it is not known how these GAPM proteins are localised across the Plasmodium life cycle, and whether different family members function alone or together. Here, we show that in two Plasmodium species GAPM2 is an IMC component whose recruitment and organisation are tightly coordinated with nuclear and cytoskeletal dynamics during parasite replication and differentiation. We find that the GAPM2 interactome remodels between asexual and sexual stages using mass spectrometry. To understand the molecular relationship between three GAPM paralogues, we solved a cryo-electron microscopy structure of the GAPM complex. This revealed an obligate heterotrimeric architecture that forms an asymmetric platform, likely to serve as a docking interface for other components of the glideosome. Finally integrating our GAPM heterotrimer structure with mass spectrometry data allowed us to propose a unified structural model of the glideosome that is conserved across apicomplexan parasites.

microbiology↗

bioRxiv2025

A divergent Plasmodium NEK4 acts as a key regulator driving the early events of meiosis

Meiosis is a conserved yet evolutionarily varied process underpinning sexual reproduction in eukaryotes. In the malaria parasite Plasmodium, meiosis is unconventional: it occurs immediately after fertilisation (post-zygotic) and must be coordinated with the transformation of the zygote into a motile ookinete. The mechanisms synchronising these meiotic and morphogenetic programmes remain unknown. Here, we identify the Plasmodium berghei NIMA-related kinase NEK4 as a key regulator that couples meiotic initiation with zygote morphogenesis. Using ultrastructure expansion microscopy, we show that NEK4 accumulates at the microtubule-organising centre (MTOC) and the apical polar complex (APC) shortly after fertilisation, preceding the assembly of perinuclear and cortical microtubules. We reveal that Plasmodium zygotes undergo MTOC-associated nuclear migration, analogous to the meiotic nuclear movement in fission yeast. Deletion of the Pbnek4 gene results in complete developmental arrest: MTOC duplication and microtubule formation are blocked, chromatin remains uncondensed, and nuclear migration and cell polarity fail to establish. Transcriptomic and phosphoproteomic analyses reveal that absence of NEK4 causes a collapse in transcriptional and phosphoregulatory networks governing meiosis and cytoskeletal organisation, leading to reduced expression and phosphorylation of important players, including HOP1, REC8, and AP2-O. These findings establish NEK4 as a key regulator driving meiotic entry and zygote maturation.

microbiology↗

bioRxiv2024

Non canonical Sun1-Allan complex orchestrates nuclear envelope remodelling and Basal Body/MTOC Segregation during rapid mitosis in Plasmodium

Mitosis in eukaryotes involves reorganization of the nuclear envelope (NE) and microtubule-organizing centres (MTOCs). During male gametogenesis in Plasmodium, the causative agent of malaria, mitosis is exceptionally rapid and highly divergent. Within 8 min, the haploid male gametocyte genome undergoes three replication cycles (1N to 8N), while maintaining an intact NE. Axonemes assemble in the cytoplasm and connect to a bipartite MTOC-containing nuclear pole (NP) and cytoplasmic basal body, producing eight flagellated gametes. The mechanisms coordinating NE remodelling, MTOC dynamics, and flagellum assembly remain poorly understood. We identify the SUN1-ALLAN complex as a novel mediator of NE remodelling and bipartite MTOC coordination during Plasmodium male gametogenesis. SUN1, a conserved NE protein, localizes to dynamic loops and focal points at the nucleoplasmic face of the spindle poles. ALLAN, a divergent allantoicase, has a location like that of SUN1, and these proteins form a unique complex, detected by live-cell imaging, ultrastructural expansion microscopy, and interactomics. Deletion of either SUN1 or ALLAN genes disrupts nuclear MTOC organization, leading to basal body mis-segregation, defective spindle assembly, and impaired spindle microtubule-kinetochore attachment, but axoneme formation remains intact. Ultrastructural analysis revealed nuclear and cytoplasmic MTOC miscoordination, producing aberrant flagellated gametes lacking nuclear material. These defects block development in the mosquito and parasite transmission, highlighting the essential functions of this complex.

cell biology↗

bioRxiv2023

Plasmodium ARK2-EB1 axis drives the unconventional spindle dynamics, scaffold formation, and chromosome segregation of sexual transmission stages

Mechanisms of cell division are remarkably diverse, suggesting the underlying molecular networks among eukaryotes differ extensively. The Aurora family of kinases orchestrates the process of chromosome segregation and cytokinesis during cell division through precise spatiotemporal regulation of their catalytic activities by distinct scaffolds. Plasmodium spp., the causative agents of malaria, are unicellular eukaryotes that have three divergent aurora-related kinases (ARKs) and lack most canonical scaffolds/activators. The parasite uses unconventional modes of chromosome segregation during endomitosis and meiosis in sexual transmission stages within mosquito host. This includes a rapid threefold genome replication from 1N to 8N with successive cycles of closed mitosis, spindle formation and chromosome segregation within eight minutes (termed male gametogony). Kinome studies had previously suggested likely essential functions for all three Plasmodium ARKs during asexual mitotic cycles; however, little is known about their location, function, or their scaffolding molecules during unconventional sexual proliferative stages. Using a combination of super-resolution microscopy, mass spectrometry, and live-cell fluorescence imaging, we set out to investigate the role of the atypical Aurora paralog ARK2 to proliferative sexual stages using rodent malaria model Plasmodium berghei. We find that ARK2 primarily localises to the spindle apparatus in the vicinity of kinetochores during both mitosis and meiosis. Interactomics and co-localisation studies reveal a unique ARK2 scaffold at the spindle including the microtubule plus end-binding protein EB1, lacking conserved Aurora scaffold proteins. Gene function studies indicate complementary functions of ARK2 and EB1 in driving endomitotic divisions and thereby parasite transmission. Our discovery of a novel Aurora kinase spindle scaffold underlines the emerging flexibility of molecular networks to rewire and drive unconventional mechanisms of chromosome segregation in the malaria parasite Plasmodium.

cell biology↗

bioRxiv2022

Cdk1-mediated threonine phosphorylation of Sam68 modulates its RNA binding, alternative splicing activity, and cellular functions

Sam68 is a member of the STAR family of proteins that directly link signal transduction with post-transcriptional gene regulation. Sam68 controls the alternative splicing of many oncogenic proteins and its role is modulated by post-translational modifications, including serine/threonine phosphorylation, that differ at various stages of the cell cycle. However, the molecular basis and mechanisms of these modulations remain largely unknown. Here, we combined mass spectrometry, NMR spectroscopy, and cell biology techniques to provide a comprehensive post-translational modification (PTM) mapping of Sam68 at different stages of the cell cycle in HEK293 and HCT116 cells. We established that Sam68 is specifically phosphorylated at T33 and T317 by Cdk1, and demonstrated that these phosphorylation events reduce the binding of Sam68 to RNA, control its cellular localization, and reduce its alternative splicing activity, leading to a reduction in the induction of apoptosis and an increase in the proliferation of HCT116 cells.

molecular biology↗

bioRxiv2021

Meta-exoproteomics identifies active plant-microbe interactions operating in the rhizosphere

The advance of DNA sequencing technologies has drastically changed our perception of the complexity and structure of the plant microbiome and its role in augmenting plant health. By comparison, our ability to accurately identify the metabolically active fraction of soil microbiota and their specific functional role is relatively limited. Here, we combined our recently developed protein extraction method and an iterative bioinformatics pipeline to enable the capture and identification of extracellular proteins (meta-exoproteomics) expressed in the rhizosphere of Brassica spp. First, we validated our method in the laboratory by successfully identifying proteins related to the host plant (Brassica rapa) and a bacterial inoculant, Pseudomonas putida BIRD-1, revealing the latter expressed numerous rhizosphere specific proteins linked to the acquisition of plant-derived nutrients. Next, we analysed natural field-soil microbial communities associated with Brassica napus L (Oil Seed rape). By combining deep-sequencing metagenomics with meta-exoproteomics, a total of 1882 proteins were identified in bulk and rhizosphere samples. Importantly, meta-exoproteomics identified a clear shift (p<0.001) in the metabolically active fraction of the soil microbiota responding to the presence of B. napus roots that was not apparent in the composition of the total microbial community (metagenome). This metabolic shift was associated with the stimulation of rhizosphere-specialised bacteria, such as Gammaproteobacteria, Betaproteobacteria and Flavobacteriia and the upregulation of plant beneficial functions related to phosphorus and nitrogen mineralisation. By providing the first meta-proteomic level assessment of the active plant microbiome at the field-scale, this study demonstrates the importance of moving past a genomic assessment of the plant microbiome in order to determine ecologically important plant: microbe interactions driving plant growth.

microbiology↗

bioRxiv2021

Activation of M1 muscarinic receptors reduce pathology and slow progression of neurodegenerative disease.

The most prevalent types of dementias, including Alzheimers disease, are those that are propagated via the spread of "prion-like" misfolded proteins. Despite considerable effort no treatments are available to slow or stop the progression of these dementias. Here we investigate the possibility that activation of the M1-muscarinic receptor (M1-receptor), which is highly expressed in the brain and that shows pro-cognitive properties, might present a novel disease modifying target. We demonstrate that the progression of murine prion disease, which we show here displays many of the pathological, behavioural and biochemical hallmarks of human neurodegenerative disease, is slowed and normal behaviour maintained by the activation of the M1-receptor with a highly tolerated positive allosteric modulator (VU846). This correlates with a reduction in both neuroinflammation and indicators of mitochondrial dysregulation, as well as a normalisation in the expression of markers associated with neurodegeneration and Alzheimers disease. Furthermore, VU846 preserves expression of synaptic proteins and post-synaptic signalling components that are altered in disease. We conclude that allosteric regulation of M1-receptors has the potential to reduce the severity of neurodegenerative diseases caused by the prion-like propagation of misfolded protein in a manner that extends life span and maintains normal behaviour.

neuroscience↗

bioRxiv2021

Impact of sulfamethoxazole on a riverine microbiome

The continued emergence of bacterial pathogens presenting antimicrobial resistance is widely recognised as a global health threat and recent attention focused on potential environmental reservoirs of antibiotic resistance genes (ARGs). Freshwater environments such as rivers represent a potential hotspot for ARGs and antibiotic resistant bacteria as they are receiving systems for effluent discharges from wastewater treatment plants (WWTPs). Effluent also contains low levels of different antimicrobials including antibiotics and biocides. Sulfonamides are antibacterial chemicals widely used in clinical, veterinary and agricultural settings and are frequently detected in sewage sludge and manure in addition to riverine ecosystems. The impact of such exposure on ARG prevalence and diversity is unknown, so the aim of this study was to investigate the release of a sub-lethal concentration of the sulfonamide compound sulfamethoxazole (SMX) on the river bacterial microbiome using a flume system. This system was a semi-natural in vitro flume using river water (30 L) and sediment with circulation to mimic river flow. A combination of omics approaches were conducted to study the impact of SMX exposure on the microbiomes within the flumes. Metagenomic analysis showed that the addition of low concentrations of SMX (<4 g L-1) had a limited effect on the bacterial resistome in the water fraction only, with no impact observed in the sediment. Metaproteomics did not show differences in ARGs expression with SMX exposure in water. Overall, the river bacterial community was resilient to short term exposure to sub-lethal concentrations of SMX which mimics the exposure such communities experience downstream of WWTPs throughout the year.

microbiology↗

bioRxiv2021

Protein Phosphatase 1 regulates atypical chromosome segregation and cell polarity during mitotic and meiotic division in Plasmodium sexual stages

PP1 is a conserved eukaryotic serine/threonine phosphatase that regulates many aspects of mitosis and meiosis, often working in concert with other phosphatases, such as CDC14 and CDC25. The proliferative stages of the malaria parasite life cycle include sexual development within the mosquito vector, with male gamete formation characterized by an atypical rapid mitosis, consisting of three rounds of DNA synthesis, successive spindle formation with clustered kinetochores, and a meiotic stage during zygote to ookinete development following fertilization. It is unclear how PP1 is involved in these unusual processes. Using real-time live-cell and ultrastructural imaging, conditional gene knockdown, RNA-seq and proteomic approaches, we show that Plasmodium PP1 is implicated in both mitotic exit and, potentially, establishing cell polarity during zygote development in the mosquito midgut, suggesting that small molecule inhibitors of PP1 should be explored for blocking parasite transmission.

cell biology↗

bioRxiv2019

Novel mechanisms for phosphate acquisition in abundant rhizosphere-dwelling Bacteroidetes

Global food production is reliant on the application of finite phosphorus (P) fertilisers. Numerous negative consequences associated with intensive P fertilisation have resulted in a high demand to find alternative sustainable methods that will enhance crop P uptake. Bacteroidetes, primarily from the genus Flavobacterium, have recently been shown to be abundant members of the plant microbiome, but their general ecological role and potential to mobilise P in the rhizosphere remains very poorly characterised. Here, we sought to determine the P mobilisation potential of Flavobacterium strains isolated from the rhizosphere of oilseed rape (Brassica napus L.). We show that these Flavobacterium strains possess novel mechanisms for P mobilisation and subsequent acquisition. These include the constitutive and inducible expression of completely novel and phylogenetically distinct phosphatases, the phosphate starvation inducible expression of uncharacterised and hypothetical genes and gene clusters and, for the first time, the involvement of outer membrane SusCD transport complexes (usually associated with carbohydrate transport) in P acquisition. The genes encoding these unusual phosphate starvation inducible proteins were enriched in plant-associated Flavobacterium strains suggesting that this machinery represents niche-adaptive strategies for overcoming P scarcity in this genus. We propose that abundant rhizosphere-dwelling Flavobacterium spp. have evolved unique mechanisms for coping with Pi-stress which may provide novel solutions for future sustainable agricultural practices.

microbiology↗