Biology subjects

Boyd, R.

Publications and source records attributed to Boyd, R..

7 recordsLinked to original sources

bioRxiv2026

Coherent Structured Illumination Microscopy with Enhanced Optical Sectioning

Coherent structured illumination microscopy (c-SIM) is a synthetic aperture optical technique for sub-diffraction limit imaging that extends the utility of traditional SIM to non-fluorescent samples. Here, we present a complementary 5-beam implementation of c-SIM that provides enhanced optical sectioning compared to conventional quadrupolar illumination. Since our approach detects intensity images due to coherent light scattering, it avoids the complications associated with detecting complex fields. Through comparative measurements on calibration samples and live microalgae, we show that 5-beam c-SIM effectively suppresses coherent defocus effects, improving image quality while simultaneously providing a 2-fold lateral resolution improvement.

bioengineering↗

bioRxiv2024

Weaker neuroligin 2-neurexin 1β interaction tethers membranes and signal synaptogenesis through clustering

Single-pass transmembrane proteins neuroligin (NL) and neurexin (NRX) constitute a pair of synaptic adhesion molecules (SAMs) that are essential for the formation of functional synapses. Binding affinities vary by [~] 1000 folds between arrays of NL and NRX subtypes, which contribute to chemical and spatial specificities. Current structures are obtained with truncated extracellular domains of NL and NRX and are limited to the higher-affinity NL1/4-NRX complexes. How NL-NRX interaction leads to functional synapses remains unknown. Here we report structures of full-length NL2 alone, and in complex with NRX1{beta} in several conformations, which has the lowest affinity among major NL-NRX subtypes. We show how conformational flexibilities may help in adapting local membrane geometry, and reveal mechanisms underlying variations in NL-NRX affinities modulation. We further show that, despite lower affinity, NL2-NRX1{beta} interaction alone is capable of tethering different lipid membranes in total reconstitution, and that NL2 and NRX1{beta} cluster at inter-cellular junctions without the need of other synaptic components. In addition, NL2 combines with the master post-synaptic scaffolding protein gephyrin and clusters neurotransmitter receptors at cellular membrane. These findings suggest dual roles of NL2 - NRX1{beta} interaction - both as mechanical tether, and as signaling receptors, to ensure correct spatial and chemical coordination between two cells to generate function synapses.

biochemistry↗

bioRxiv2024

The role of phosphorylation and dephosphorylation in the regulation of Rubisco activase

Rubisco activase is an ATP-dependent chaperone that facilitates dissociation of inhibitory sugar phosphates from the catalytic sites of ribulose-1,5-bisphosphate carboxylase/oxygenase during photosynthesis. In Arabidopsis, Rubisco activase is negatively regulated by dark-dependent phosphorylation of threonine 78. The prevalence of threonine 78 in Rubisco activase was investigated across sequences from 91 plant species, finding 29 ([~]32%) species shared a threonine in the same position. Analysis of seven C3 species with an antibody raised against a threonine 78 phospho-peptide demonstrated that this position is phosphorylated in multiple genera. However, light-dependent dephosphorylation of threonine 78 was observed only in Arabidopsis. Further, phosphorylation of threonine 78 could not be detected in any of the four C4 grass species examined. The results suggest that despite conservation of threonine 78 in Rubisco activase from a wide range of species, a regulatory role for phosphorylation at this site is more limited. This provides a case study for how variation in post-translational regulation can amplify functional divergence across the phylogeny of plants beyond what is explained by sequence variation in a metabolically important protein.

plant biology↗

bioRxiv2024

The refinement paradox and cumulative cultural evolution: collective improvement in knowledge favors conformity, blind copying and hyper-credulity

Social learning is common in nature, yet cumulative culture (where knowledge and technology increase in complexity and diversity over time) appears restricted to humans. To understand why, we organized a computer tournament in which programmed entries specified when to learn new knowledge and when to refine (i.e. improve) existing knowledge. The tournament revealed a refinement paradox: refined behavior afforded higher payoffs as individuals converged on a small number of successful behavioral variants, but refining did not generally pay. Paradoxically, entries that refined only in certain conditions did best during behavioral improvement, while simple copying entries thrived when refinement levels were high. Cumulative cultural evolution may be rare in part because sophisticated strategies for improving knowledge and technology are initially advantageous, yet complex culture, once achieved, favors conformity, blind imitation and hyper-credulity.

evolutionary biology↗

bioRxiv2021

Mapping interindividual dynamics of innate immune response at single-cell resolution

Common genetic variants modulate the cellular response to viruses and are implicated in a range of immune pathologies, including infectious and autoimmune diseases. The transcriptional antiviral response is known to vary between infected cells from a single individual, yet how genetic variants across individuals modulate the antiviral response (and its cell-to-cell variability) is not well understood. Here, we triggered the antiviral response in human fibroblasts from 68 healthy donors, and profiled tens of thousands of cells using single-cell RNA-seq. We developed GASPACHO (GAuSsian Processes for Association mapping leveraging Cell HeterOgeneity), the first statistical approach designed to identify dynamic eQTLs across a transcriptional trajectory of cell populations, without aggregating single-cell data into pseudo-bulk. This allows us to uncover the underlying architecture and variability of antiviral response across responding cells, and to identify more than two thousands eQTLs modulating the dynamic changes during this response. Many of these eQTLs colocalise with risk loci identified in GWAS of infectious and autoimmune diseases. As a case study, we focus on a COVID-19 susceptibility locus, colocalised with the antiviral OAS1 splicing QTL. We validated it in blood cells from a patient cohort and in the infected nasal cells of a patient with the risk allele, demonstrating the utility of GASPACHO to fine-map and functionally characterise a genetic locus. In summary, our novel analytical approach provides a new framework for delineation of the genetic variants that shape a wide spectrum of transcriptional responses at single-cell resolution.

genetics↗

bioRxiv2019

Brodifacoum does not modulate human cannabinoid receptor-mediated hyperpolarization of AtT20 cells or inhibition of adenylyl cyclase in HEK 293 cells

BackgroundSynthetic cannabinoids are a commonly used class of recreational drugs that can have significant adverse effects. There have been sporadic reports of co-consumption of illicit drugs with rodenticides such as warfarin and brodifacoum (BFC) over the past 20 years but recently, hundreds of people have been reported to have been poisoned with a mixture of synthetic cannabinoids and BFC. We have sought to establish whether BFC directly affects cannabinoid receptors, or their activation by the synthetic cannabinoid CP55940 or the phytocannabinoid {Delta}9-tetrahydrocannabinol ({Delta}9-THC).\n\nMethodsThe effects of BFC on the hyperpolarization of wild type AtT20 cells, or AtT20 cells stably expressing human CB1- and CB2-mediated receptors, were studied using a fluorescent assay of membrane potential. The effects of BFC on CB1 and CB2 mediated inhibition of forskolin-stimulated adenylyl cyclase (AC) activation was measured using a BRET assay of cAMP levels in HEK 293 cells stably expressing human CB1 and CB2.\n\nResultsBFC did not activate CB1 or CB2 receptors, or affect the hyperpolarization of wild type AtT20 cells produced by somatostatin. BFC (10 {micro}M) did not affect the hyperpolarization of AtT20-CB1 or AtT20-CB2 cells produced by CP55940 or {Delta}9-THC. BFC (1 {micro}M) did not affect the inhibition of forskolin-stimulated AC activity by CP55940 in HEK 293 cells expressing CB1 or CB2. BFC (1 {micro}M) also failed to affect the desensitization of CB1 and CB2 signalling produced by prolonged (30 min) application of CP55940 or {Delta}9-THC to AtT20 cells.\n\nDiscussionBFC is not a cannabinoid receptor agonist, and appeared not to affect cannabinoid receptor activation. Our data suggests there is no pharmacodynamic rationale for mixing BFC with synthetic cannabinoids, however, it does not speak to whether BFC may affect synthetic cannabinoid metabolism or biodistribution. The reasons underlying the mixing of BFC with synthetic cannabinoids are unknown, and it remains to be established whether the \"contamination\" was deliberate or accidental. However, the consequences for people who ingested the mixture were often serious, and sometimes fatal, but this seems unlikely to be due to BFC action at cannabinoid receptors.

pharmacology and toxicology↗

bioRxiv2018

Temperature response of Rubisco kinetics in Arabidopsis thaliana: thermal breakpoints and implications for reaction mechanisms

Optimization of Rubisco kinetics could improve photosynthetic efficiency, ultimatly resulting in increased crop yield. However, imprecise knowledge of the reaction mechanism and the individual rate constants limit our ability to optimize the enzyme. Membrane inlet mass spectrometery (MIMS) may offer benefits over traditional methods for determining individual rate constants of the Rubisco reaction mechanism, as it can directly monitor concentration changes in CO2, O2, and their isotopologs during assays. However, a direct comparsion of MIMS to the traditional Radiolabel method of determining Rubisco kinetic parameters has not been made. Here, the temperature responses of Rubisco kinetic parameters from Arabidopsis thaliana were measured using the Radiolabel and MIMS methods. The two methods provided comparable parameters above 25 {degrees}C, but temperature responses deviated at low temperature as MIMS derived catalytic rates of carboxylation, oxygenation, and CO2/O2 specificity showed thermal breakpoints. Here we discuss the variability and uncertainty surrounding breakpoints in the Rubisco temperature response and relavance of individual rate constants of the reaction mechanisms to potential breakpoints.

plant biology↗