Biology subjects

Buckley, D.

Publications and source records attributed to Buckley, D..

5 recordsLinked to original sources

bioRxiv2025

Molecular insights into electroreceptor ribbon synapses from differential gene expression in sturgeon lateral line organs

In fishes and aquatic-stage amphibians, mechanosensory neuromasts are arranged in characteristic lines in the skin of the head and trunk, with afferent innervation from anterior or posterior lateral line nerves. In electroreceptive non-teleost jawed fishes and amphibians, fields of electrosensory ampullary organs flank some or all of the cranial neuromast lines, innervated by the anterior lateral line nerve. Like the mechanosensory hair cells found in neuromasts and the inner ear, electroreceptor cells in ampullary organs form specialised ribbon synapses with afferent nerve terminals. Ribbon synapses in hair cells are distinct from other glutamatergic synapses, including the ribbon synapses in photoreceptors: otoferlin is the Ca2+ sensor for synaptic vesicle exocytosis and synaptic vesicles are loaded with glutamate by vGlut3. We previously showed that the genes encoding otoferlin and vGlut3 are expressed by ampullary organs as well as neuromasts in a chondrostean ray-finned fish, the Mississippi paddlefish (Polyodon spathula), suggesting that electroreceptor ribbon synapses are very similar to those in hair cells. In this study, we selected seven additional synapse-related candidate genes from our previously published dataset of putatively lateral line organ-enriched genes from late-larval paddlefish, and examined their expression in developing lateral line organs in a related chondrostean, the sterlet sturgeon (Acipenser ruthenus). We found that genes encoding the presynaptic cell adhesion molecule Nrxn3, the calcium-independent synaptotagmin Syt14, the high-affinity glutamate re-uptake transporter EAAT1 (GLAST), calmodulin regulator protein PCP4 (PEP-19) and cell adhesion molecule DSCAML1 were expressed in both neuromasts and ampullary organs. In contrast, Cbln18, encoding a secreted trans-synaptic scaffolding protein, was only expressed in neuromasts and Tulp1, encoding tubby-related protein 1 (required for the development and function of photoreceptor ribbon synapses), was only expressed in ampullary organs. Our results support electroreceptor ribbon synapses being glutamatergic and suggest further commonalities, but also some differences, with hair cell ribbon synapses.

developmental biology↗

bioRxiv2025

Parenteral Vaccination with recombinant EtpA glycoprotein impairs enterotoxigenic E. coli colonization

Enterotoxigenic E. coli (ETEC) cause hundreds of millions of cases of acute diarrheal illness in low-middle income regions, disproportionately in young children. To date there is no licensed, broadly protective vaccine to protect against these common but antigenically heterogeneous pathogens. One of the more highly conserved antigens of ETEC, EtpA, is an extracellular glycoprotein adhesin that preferentially binds to blood group A glycans on intestinal epithelia. EtpA contributes to increased severity of illness in blood group A individuals, elicits robust serologic and fecal antibody responses following infection, and has been associated with protection against subsequent infection. However, its utility as a protective antigen needs further examination. In the present studies we examined whether parenteral vaccination with recombinant EtpA (rEtpA) could afford protection against intestinal colonization in a murine model of ETEC infection. Here, we demonstrate that intramuscular vaccination with rEtpA when adjuvanted with double mutant LT (dmLT) primes IgG predominant mucosal antibody responses to ETEC challenge. Notably, however, both antibody levels and avidity, as well as protection were dependent on vaccination schedule. Likewise, by electron microscopy polyclonal epitope mapping (EMPEM) we observed a greater diversity of epitopes targeted by antibodies after a more protracted vaccination schedule. Next, we explored the utility of IM immunization with alum-adjuvanted rEtpA. This elicited strong serologic and fecal IgG responses. Although accompanied by negligible IgA mucosal responses, EtpA alum-adjuvanted IM vaccination nevertheless protected against ETEC intestinal colonization. Collectively, these data suggest that EtpA could expand the portfolio of antigens targeted in ETEC subunit vaccine development.

microbiology↗

bioRxiv2024

A small molecule VHL molecular glue degrader for cysteine dioxygenase 1

The Von Hippel-Lindau Tumor Suppressor gene product (pVHL) is an E3 ligase substrate receptor that binds proline-hydroxylated HIF1-, leading to its ubiquitin-dependent degradation. By using protein arrays, we identified a small molecule that binds the HIF1- binding pocket on pVHL and functions as a molecular glue degrader of the neosubstrate cysteine dioxygenase (CDO1) by recruiting it into the VHL-cullin-ring E3 ligase complex and leading to its selective degradation. The CDO1 binding region involved in VHL recruitment was characterized through a combination of mutagenesis and protein-protein docking coupled with molecular dynamics-based solvation analysis. The X-ray structure of the ternary complexes of VHL, CDO1, and degrader molecules confirms the binding region prediction and provides atomic insights into key molecular glue interactions.

biochemistry↗

bioRxiv2020

Retinotopic remapping in the visual system of deaf adults

Deaf individuals rely on visual rather than auditory cues to detect events in the periphery, putting a greater demand on neural resources for vision. Comparing visual maps in the brains of early deaf and hearing adults, we found a redistribution of neural resources in the lateral geniculate nucleus and primary visual cortex, with larger representations of the periphery, at a cost of smaller representations of the central visual field.

neuroscience↗

bioRxiv2018

Brief Freezing Steps Lead to Robust Immunofluorescence in Drosophila Larval Brains

Drosophila melanogaster possess complex neuronal networks regulating sophisticated behavioral outputs that aid in studying the molecular mechanisms of neuronal function and neurodegenerative disease. Immunofluorescence (IF) techniques provide a way to visualize the spatiotemporal organization of these networks, permitting observation of their development, functional location, remodeling, and eventually - degradation. However, general immunostaining techniques do not always result in sufficient antibody penetration through the brain, and techniques used to enhance permeability can compromise structural integrity. We have found that freezing larval brains facilitates permeability with no apparent loss of antibody specificity or structural integrity. To demonstrate the advantage of this freezing technique, we compared results to two commonly used permeation methods: Detergent alone (Basic) and proteolytic degradation (Collagenase) techniques.\n\nSummaryHere we compare four different immunofluorescence techniques demonstrating that freezing Drosophila brains results in robust staining of small neurons in the larval brain without compromising structural integrity.

cell biology↗