Biology subjects

Beard, M.

Publications and source records attributed to Beard, M..

3 recordsLinked to original sources

bioRxiv2023

Botulinum toxin intoxication requires retrograde transport and membrane translocation at the ER

Botulinum neurotoxin A (BoNT/A) is a highly potent proteolytic toxin specific for neurons with numerous clinical and cosmetic uses. After uptake at the synapse, the protein is proposed to translocate from synaptic vesicles to cytosol through a self-formed channel. Surprisingly, we found that after intoxication proteolysis of a fluorescent reporter occurs in the neuron soma first and then centrifugally in neurites. To investigate the molecular mechanisms at play, we use a genome-wide siRNA screen in genetically engineered neurons and identify over three hundred genes. An organelle-specific split-mNG complementation indicates BoNT/A traffic from the synapse to the soma-localised Golgi in a retromer dependent fashion. The toxin then moves to the ER and appears to require the Sec61 complex for retro-translocation to the cytosol. Our study identifies genes and trafficking processes hijacked by BoNT/A, revealing a complex route for efficient intoxication that contradicts the currently accepted model of BonT intoxication.

cell biology↗

bioRxiv2023

Structure and activity of botulinum neurotoxin X

Botulinum neurotoxins (BoNTs) are the most potent toxins known and are used to treat an increasing number of medical disorders. All BoNTs are naturally co-expressed with a protective partner protein (NTNH) with which they form a 300 kDa complex, to resist acidic and proteolytic attack from the digestive tract. We have previously identified a new botulinum neurotoxin serotype, BoNT/X, that has unique and therapeutically attractive properties. We present the cryo-EM structure of the BoNT/X-NTNH/X complex at 3.1 [A] resolution. Unexpectedly, the BoNT/X complex is stable and protease resistant at both neutral and acidic pH and disassembles only in alkaline conditions. Using the stabilizing effect of NTNH, we isolated BoNT/X and showed that it has very low potency both in vitro and in vivo. Given the high catalytic activity and translocation efficacy of BoNT/X, low activity of the full toxin is likely due to the receptor-binding domain, which presents weak ganglioside binding and exposed hydrophobic surfaces.

biochemistry↗

bioRxiv2018

The Antiviral Protein Viperin Enhances The Host Interferon Response Following STING Activation

Viperin is an interferon-inducible protein that is pivotal for eliciting an effective immune response against an array of diverse viral pathogens. Here we describe a mechanism of viperins broad antiviral activity by demonstrating the proteins ability to synergistically enhance the innate immune dsDNA signalling pathway to limit viral infection. Viperin co-localised with the key signalling molecules of the innate immune dsDNA sensing pathway, STING and TBK1; binding directly to STING and inducing enhanced K63-linked polyubiquitination of TBK1. Subsequent analysis identified viperins necessity to bind the cytosolic iron-sulphur assembly component 2A, to prolong its enhancement of the type-I interferon response to aberrant dsDNA. Here we show that viperin facilitates the formation of a signalling enhanceosome, to coordinate efficient signal transduction following activation of the dsDNA signalling pathway; which results in an enhanced antiviral state. We also provide evidence for viperins radical SAM enzymatic activity to self-limit its immunomodulatory functions. This data further defines viperins role as a positive regulator of innate immune signalling, offering a mechanism of viperins broad antiviral capacity.

immunology↗