Biology subjects

Dillon, S.

Publications and source records attributed to Dillon, S..

11 recordsLinked to original sources

bioRxiv2026

TNAP and PHOSPHO1 function synergistically to afford critical control over the mineralisation of the postnatal murine skeleton

Biomineralisation is essential for skeletal integrity, yet the synergistic roles of tissue non-specific alkaline phosphatase (TNAP) and PHOSPHO1 in postnatal bone mineralisation remain poorly defined. To decipher this, we generated a novel murine model in which Alpl was deleted in Prx1- expressing cells (AlplPrx1/Prx1) in mice with a global Phospho1-/- deficiency to overcome the perinatal lethality that arises upon dual global deletion. Using a multi-modal approach to spatially phenotype the limbs of these animals, we reveal mice lacking both TNAP and PHOSPHO1 exhibit a distinct lack of mineralisation and altered anatomical structure at postnatal day 1 (PN1) and 3-weeks of age. Although viable, these mice did not thrive due to their reduced size, thus further investigations were conducted on mice with a heterozygous deletion of TNAP (Alplwt/Prx1;Phospho1-/-). Although smaller than wild-types at PN1 and 3 weeks old, these mice did not display the gross limb deformations observed in the homozygous animals and the single, functioning Alpl allele rescued the loss of biomineralisation observed following dual phosphatase deletion. At 6-weeks of age, compromised epiphyses and metaphyses were only seen in AlplPrx1/Prx1 animals. Further, we found that tibial geometry and porosity was significantly altered by Phospho1 deletion (Phospho1-/-), which was compounded in the Alplwt/Prx1;Phospho1-/- mice and linked to alterations in collagen configuration, matrix mineralisation and growth plate deformities. Together, our findings establish the mechanistic framework for TNAP and PHOSPHO1 in permissive biomineralisation, providing critical insights into this fundamental process. Significance StatementBiomineralisation is essential for skeletal development and is critically dependent on phosphatases that release inorganic phosphate for hydroxyapatite formation. Our study investigates the dual role of PHOSPHO1 and TNAP in this process, using a novel murine knockout model. Deletion of both enzymes results in complete loss of bone mineralisation, demonstrating their critical synergistic function. Further, we show that PHOSPHO1 and TNAP exhibit distinct, spatially-restricted functions in the tibia and thus enhances our understanding of the fundamentals processes underpinning biomineralisation. These findings also have clinical relevance as they have the potential to inform on treatment strategies for hypo- and hyper-mineralised pathologies.

developmental biology↗

bioRxiv2025

Inorganic pyrophosphate disrupts amorphous hydrated bone mineral interfaces in hypophosphatasia

Bone mineral molecular architecture is tightly regulated by the kinetics of calcium phosphate phase transformations. In the rare skeletal disease hypophosphatasia (HPP), caused by inactivating mutations in the ALPL gene encoding tissue-nonspecific alkaline phosphatase (TNSALP), accumulation of inorganic pyrophosphate (PPi) alters these phase dynamics. Using solid-state nuclear magnetic resonance spectroscopy and high-resolution electron microscopy, in patient samples we show that bone mineral from compound heterozygous HPP patients exhibits a loss of hydrated amorphous interfacial phases and instead contains highly crystalline hydroxyapatite (HAP), correlating with abnormally high bone mineral density and brittle atypical femoral fractures. Synthetic and cellular models demonstrate that elevated PPi impedes normal phase transitions from amorphous calcium phosphate precursors, bypassing intermediate states and driving ordered HAP nucleation. These findings link disrupted mineral phase kinetics to pathological bone mineral molecular structure, emphasising the critical importance of the hydrated amorphous shell around bone mineral for its material properties and redefining HPP as a molecular mineralization disorder. Our findings also illustrate how biochemical cues regulate non-equilibrium crystallization pathways in biomineralization.

biophysics↗

bioRxiv2025

Proteomic characterisation of matrix vesicles from primary osteoblasts indicates a mixed population with both exosomal and ectosomal routes of biogenesis.

Matrix Vesicles are a crucial step in producing a mineralised, healthy skeleton. Released from chondrocytes and osteoblasts, they concentrate calcium and phosphate to establish the deposition of hydroxyapatite around and within the collagen fibrils of the extracellular matrix, becoming embedded in the matrix in the process. In the 55 years since H. Clarke Anderson first described them, blebbing from the surface of chondrocytes, a consensus on their role, contents and their biogenesis, has yet to be reached. This is in part due to the range of cell types from which they are released and the multitude techniques that can be employed to isolate them. In this study, we, for the first time, characterise the proteome of matrix vesicles isolated from primary osteoblasts. By focussing on those vesicles that have become embedded within the matrix, we are able to avoid bias for specific modes of biogenesis and focus only on osteoblast-released vesicles that are associated with the matrix. Moreover, by studying these vesicles over a time course of mineralising activity, we are able to identify changes in the properties of these vesicles, and develop a more accurate picture of which proteins are involved specifically in mineralisation. In particular we identify the presence of markers associated with the ectosomal and exosomal release of matrix vesicles, as well as identifying proteins required for the maintenance and mineralisation of the extracellular matrix. These data portray a heterogeneous population of matrix vesicles, with different roles to play in bone development.

cell biology↗

bioRxiv2025

Advancing rust resistance in elite wheat with haplotype mapping and a novel introgression strategy

Wheat production is continually threatened by stripe and leaf rust because virulent races rapidly overcome single race-specific genes. Durable, broad-spectrum resistance is needed. Adult plant resistance (APR) provides partial, stable resistance from multiple minor-effect loci acting additively; pleiotropic loci like Lr34/Yr18 and Lr46/Yr29 add durability. We used an elite Australian panel (OzWheat=589) and a diverse landrace panel (Vavilov=295), genotyped with [~]30K SNPs and phenotyped across environments. Linkage disequilibrium partitioning defined 7,659 genome-wide haploblocks. To prioritise robust signals, we ranked haploblocks by haplotype effect variance and examined the top 100 per trait. For stripe rust, 52/100 were significant, with 32 shared across panels; for leaf rust, 50 were significant, 29 also detected in Vavilov. Several intervals co-localised with APR regions (Lr46/Yr29), and one 7BL interval intersected seedling gene Lr14a. To translate mapping into breeding decisions, we developed an introgression fitness index to quantify the value of resistant haplotypes in elite backgrounds. Using elite cultivar Scepter, we applied a genetic algorithm to select 50 donor parents carrying desirable haplotypes. Simulations showed that pyramiding these haplotypes can enhance resistance while maintaining elite genomic background. This study provides practical breeding tools, including haplotype catalogue and a novel selection index to accelerate rust-resistant wheat development.

genetics↗

bioRxiv2025

Lysyl oxidase-mediated intermolecular crosslinks fine-tune collagen I molecular dynamics and regulate cell-matrix interactions through focal adhesions

Lysyl oxidase (LOX)-mediated intermolecular crosslinking is essential for collagen I fibril stability, yet its influence on collagen molecular conformation and dynamics, and the downstream consequences for cell-matrix interactions remain poorly understood. Here, we genetically modulated LOX in collagen I-producing MC3T3-E1 cells to generate matrices with elevated (overexpression, OX) or absent (knockout, KO) crosslinking. Enhanced crosslinking yielded thick, continuous, aligned fibrils, whereas reduced crosslinking produced friable, dissociated fibrils. Solid-state nuclear magnetic resonance spectroscopy (SSNMR) revealed local triple-helix unfolding and altered nanosecond- and microsecond-scale molecular motions in both OX and KO matrices, changes largely reversible upon decellularization, implicating a synergistic role of crosslinking chemistry and cell-applied forces in regulating the dynamically-accessible conformations of collagen I. Changes in the molecular structure and dynamics of collagen had a functional impact on cell adhesion and mechanotransduction. These findings identify collagen crosslinking as a tunable element of the extracellular matrix "mechanical code," integrating biochemical modification with molecular-scale mechanics to regulate cell-matrix adhesion and mechanosignalling.

biophysics↗

bioRxiv2025

Human Peripheral Nerve-on-a-Chip on a Multiwell Microelectrode Array as a Scalable Preclinical Neurotoxicity Assay

Reliable human-relevant models of peripheral nerve function remain a critical unmet need in preclinical drug development, particularly for predicting neurotoxicity and bridging the gap to clinical translation. Here, we introduce a next generation Nerve-on-a-Chip, PNS-3D organoids, as a novel human-cell-based 3D peripheral nerve microphysiological system (MPS) that recapitulates key functional and structural features of native nerves, including long-distance axonal outgrowth, physiological myelination, and clinically translational population level electrophysiology. The platform integrates iPSC-derived human sensory neurons and primary human Schwann cells within a spatially organized 3D environment, coupled to a custom embedded electrode array that enables high-content, longitudinal, and clinically translatable functional assessments. As a proof of concept, we evaluated the platforms predictive power using vincristine, a chemotherapeutic agent known to cause chemotherapy-induced peripheral neuropathy (CIPN). PNS-3D organoids captured dose- and time-dependent deficits in nerve conduction velocity, compound action potential amplitude, and axonal degeneration, with IC values in line with human clinical exposures-- outperforming traditional 2D cultures and in vivo benchmarks. Transcriptomic and morphological analyses further revealed neuron-specific degeneration consistent with axonopathy. These results validate the platform as a human-relevant, clinically translation, and scalable solution, enabling mechanistic safety assessment and drug discovery for neurotoxic and neuroprotective therapeutics.

pharmacology and toxicology↗

bioRxiv2024

Incorporating gene expression and environment improves genomic prediction of wheat traits

The adoption of novel molecular strategies such as genomic selection (GS) in crop breeding have been key to maintaining rates of genetic gain through increased efficiency and shortening the cycle of evaluation relative to conventional selection. In the search for improved methodologies that incorporate novel sources of variation for the assessment of genetic merit, GS remains a focus of crop breeding research globally. Here we explored the role transcrip-tome data could play in enhancing GS using wheat as a test case. Across 286 wheat lines, we integrated phenotype and multi-omic data from controlled environment and field experiments including ca. 40K single nucleotide polymorphisms (SNP), abundance data for ca. 50K transcripts as well as meta-data (e.g. categorical environments) predicted individual genetic merit for two agronomic traits, flowering time and height. We combined phenotype and multi-omic data from both controlled environments and field experiments. This included ca. 40K single nucleotide polymorphisms (SNPs), ca. 50K transcript abundance data, and metadata (such as categorical environmental conditions). Using this integrated data, we predicted individual genetic merit for two agronomic traits: flowering time and height. We evaluated the performance of different model scenarios based on linear (GBLUP) and Gaussian/nonlinear (RKHS) regression in the Bayesian analytical frame-work. These models explored the relative contributions of different combinations of explanatory variables; additive genomic (G), transcriptomic (T) and environment (E), with and without considering non-additive epistasis and the GxE random effects. In controlled environments, where traits were measured under contrasting daylength regimes (long and short days), transcriptome abundance outperformed other explanatory variables when considered independently, while the model combining SNP, environment and G x E marginally outperformed the transcriptome. The best performing model for prediction of both flowering and height combined all data types, G x E and epistasis, where the GBLUP framework showed slightly better performance overall compared with RKHS across all tests. Under field conditions, we similarly found that models combining all variables were superior, with the GBLUP and RKSH methods performing equally well. However, the relative contribution of the transcriptome was reduced. Our results show there is a predictive advantage to direct inclusion of the transcriptome for genomic evaluation in wheat breeding. However, the complexity and cost of generating large scale transcriptome data are likely to limit its feasibility for commercial breeding. We demonstrate that combining less costly environmental covariates with conventional genomic data provide a practical alternative with similar gains to the transcriptome when environments are well characterised. HighlightsO_LIIncorporating transcriptome and environment in genomic prediction; C_LIO_LIModel comparisons and Bayesian inference. C_LIO_LIDifferential random effects of transcriptome, SNP and environment. C_LI

genomics↗

bioRxiv2023

Mitochondrial dysfunction and mitophagy blockade contribute to renal osteodystrophy in chronic kidney disease-mineral bone disorder

Chronic kidney disease-mineral and bone disorder (CKD-MBD) presents with extra-skeletal calcification and renal osteodystrophy (ROD). The origins of ROD likely lie with elevated uremic toxins and/or an altered hormonal profile but the cellular events responsible remain unclear. Here, we report that stalled mitophagy contributes to mitochondrial dysfunction in bones of a CKD-MBD mouse model, and also human CKD-MBD patients. RNA-seq analysis exposed an altered expression of genes associated with mitophagy and mitochondrial function in tibia of CKD-MBD mice. The accumulation of damaged osteocyte mitochondria and the expression of mitophagy regulators, p62/SQSTM1, ATG7 and LC3 was inconsistent with functional mitophagy, and in mito-QC reporter mice with CKD-MBD, there was a 2.3-fold increase in osteocyte mitolysosomes. Altered expression of mitophagy regulators in human CKD-MBD bones was also observed. To determine if uremic toxins were possibly responsible for these observations, indoxyl sulfate treatment of osteoblasts revealed mitochondria with distorted morphology and whose membrane potential and oxidative phosphorylation were decreased, and oxygen-free radical production increased. The altered p62/SQSTM1 and LC3-II expression was consistent with impaired mitophagy machinery and the effects of indoxyl sulfate were reversible by rapamycin. In conclusion, mitolysosome accumulation from impaired clearance of damaged mitochondria may contribute to the skeletal complications, characteristic of ROD. Targeting mitochondria and the mitophagy process may therefore offer novel routes for intervention to preserve bone health in patients with ROD. Such approaches would be timely as our current armamentarium of anti-fracture medications has not been developed for, or adequately studied in, patients with severe CKD-MBD. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=113 SRC="FIGDIR/small/573355v1_ufig1.gif" ALT="Figure 1"> View larger version (51K): org.highwire.dtl.DTLVardef@1f23caforg.highwire.dtl.DTLVardef@6e3e94org.highwire.dtl.DTLVardef@1ce6adeorg.highwire.dtl.DTLVardef@ca5558_HPS_FORMAT_FIGEXP M_FIG C_FIG TRANSLATIONAL STATEMENTRenal osteodystrophy (ROD) remains the major skeletal complication of chronic kidney disease-mineral and bone disorder (CKD-MBD). As a disease characterised by biochemical and hormone abnormalities, ROD is exacerbated by osteocyte mitochondrial dysfunction. Advances in our understanding of the mitophagy pathway are vital to improving the clinical management of ROD. The dysregulation of mitophagy in murine and human CKD-MBD bone provided evidence of delayed clearance of damaged mitochondria, which was also observed in uremic toxin-treated-osteoblasts but reversible upon rapamycin treatment. This study reveals the therapeutic potential of managing ROD by restoring defective mitophagy in osteocytes.

genomics↗

bioRxiv2023

Microbiota-dependent indole production is required for the development of collagen-induced arthritis

Altered tryptophan catabolism has been identified in inflammatory diseases like rheumatoid arthritis (RA) and spondyloarthritis (SpA), but the causal mechanisms linking tryptophan metabolites to disease are unknown. Using the collagen-induced arthritis (CIA) model we identify alterations in tryptophan metabolism, and specifically indole, that correlate with disease. We demonstrate that both bacteria and dietary tryptophan are required for disease, and indole supplementation is sufficient to induce disease in their absence. When mice with CIA on a low-tryptophan diet were supplemented with indole, we observed significant increases in serum IL-6, TNF, and IL-1{beta}; splenic ROR{gamma}t+CD4+ T cells and ex vivo collagen-stimulated IL-17 production; and a pattern of anti-collagen antibody isotype switching and glycosylation that corresponded with increased complement fixation. IL-23 neutralization reduced disease severity in indole-induced CIA. Finally, exposure of human colon lymphocytes to indole increased expression of genes involved in IL-17 signaling and plasma cell activation. Altogether, we propose a mechanism by which intestinal dysbiosis during inflammatory arthritis results in altered tryptophan catabolism, leading to indole stimulation of arthritis development. Blockade of indole generation may present a novel therapeutic pathway for RA and SpA.

immunology↗

bioRxiv2021

Differential Expression of Alkaline Phosphatase and PHOSPHO1 in Bone from a Murine Model of Chronic Kidney Disease

Patients with advanced chronic kidney disease (CKD) often present with skeletal abnormalities; a condition known as renal osteodystrophy (ROD). While Tissue-nonspecific alkaline phosphatase (TNAP) and PHOSPHO1 are recognized to be critical for bone mineralization, their role in the etiology of ROD is unclear. To address this, ROD was induced in both wild-type and Phospho1 knockout (P1KO) mice using dietary adenine supplementation. The mice presented with hyperphosphatemia, hyperparathyroidism, and elevated levels of FGF23 and bone turnover markers. In particular, we noted that in CKD mice, bone mineral density (BMD) was increased in cortical bone (p < 0.05) but decreased in trabecular bone (p < 0.05). These changes were accompanied by decreased TNAP (p < 0.01) and increased PHOSPHO1 (p < 0.001) expression in wild-type CKD bones. In P1KO CKD mice, the cortical BMD phenotype was rescued, suggesting that the increased cortical BMD of CKD mice was driven by increased PHOSPHO1 expression. Other structural parameters were also improved in P1KO CKD mice. We further investigated the driver of the mineralization defects, by studying the effects of FGF23, PTH, and phosphate administration on PHOSPHO1 and TNAP expression by primary murine osteoblasts. We found both PHOSPHO1 and TNAP expression to be down-regulated in response to phosphate and PTH. While matrix mineralization was increased with phosphate (Pi), it decreased with PTH and FGF23 had no effect. The in vitro data suggest that the TNAP reduction in CKD-MBD is driven by the hyperphosphatemia and/or hyperparathyroidism noted in these mice, while the higher PHOSPHO1 expression may be a compensatory mechanism in an attempt to protect the bone from hypomineralization. We propose that increased PHOSPHO1 expression in ROD may contribute to the disordered skeletal mineralization characteristic of this progressive disorder.

cell biology↗

bioRxiv2021

FXR inhibition reduces ACE2 expression, SARS-CoV-2 infection and may improve COVID-19 outcome

Prevention of SARS-CoV-2 entry in cells through the modulation of viral host receptors, such as ACE2, could represent a new therapeutic approach complementing vaccination. However, the mechanisms controlling ACE2 expression remain elusive. Here, we identify the farnesoid X receptor (FXR) as a direct regulator of ACE2 transcription in multiple COVID19-affected tissues, including the gastrointestinal and respiratory systems. We demonstrate that FXR antagonists, including the over-the-counter compound z-guggulsterone (ZGG) and the off-patent drug ursodeoxycholic acid (UDCA), downregulate ACE2 levels, and reduce susceptibility to SARS-CoV-2 infection in lung, cholangiocyte and gut organoids. We then show that therapeutic levels of UDCA downregulate ACE2 in human organs perfused ex situ and reduce SARS-CoV-2 infection ex vivo. Finally, we perform a retrospective study using registry data and identify a correlation between UDCA treatment and positive clinical outcomes following SARS-CoV-2 infection, including hospitalisation, ICU admission and death. In conclusion, we identify a novel function of FXR in controlling ACE2 expression and provide evidence that this approach could be beneficial for reducing SARS-CoV-2 infection, thereby paving the road for future clinical trials.

microbiology↗