Biology subjects

White, I. J.

Publications and source records attributed to White, I. J..

9 recordsLinked to original sources

bioRxiv2026

PKA can adopt structural or enzymatic roles depending on its anchoring site

Protein kinase A (PKA) is a central mediator of cAMP signaling whose spatial specificity is conferred by A-kinase anchoring proteins (AKAPs). However, functions of individual anchored PKA pools are difficult to resolve as current approaches cannot isolate anchoring at defined sites. Here, we develop a SpyCatcher-SpyTag-based replacement strategy enabling direct comparison of AKAP function with and without type II PKA regulatory (RII) subunit anchoring in intact signaling environments. Anchoring RII to AKAP79 or AKAP1 produced distinct phosphorylation-dependent effects, including catalytic subunit membrane recruitment and mitochondrial morphology regulation. In contrast, MAP2-anchored RII drove phosphorylation-independent microtubule straightening, increased formation of widely spaced bundles, and supported dendritic arborization. This structural function was recapitulated by native MAP2-RII binding and required an intact cAMP-binding pocket in the second cyclic nucleotide-binding domain. These findings establish that AKAP-bound PKA can function as either an enzymatic or structural component and provide a generalizable strategy to dissect compartmentalized kinase signaling.

molecular biology↗

bioRxiv2026

Gut-relevant short-chain fatty acids modulate host-pathogen dynamics of uropathogenic Escherichia coli at the colonic epithelial interface

Urinary tract infection (UTI) ranks among the most prevalent bacterial infections worldwide, affecting over 400 million people each year. Uropathogenic Escherichia coli (UPEC), the main aetiological cause of UTI, colonises the intestinal tract, which is thought to serve as a distal reservoir for gut-UTI recurrence. Despite this, the precise role of the gut in UTI recurrence is still not fully defined. Recent research investigating the gut-UTI axis has revealed that reduced abundance of gut commensals producing short-chain fatty acids (SCFAs, namely acetate, butyrate and propionate) is associated with recurrent and chronic UTI. We therefore aimed to investigate the impact of these gut commensal-derived metabolites on a diverse panel of UPEC strains, including well-studied prototypical strains (UTI89, CFT073), a non-pathogenic isolate E. coli K-12, and various clinical UTI isolates (from the urine of both symptomatic and asymptomatic individuals). We observed that SCFAs modulate bacterial growth kinetics in a concentration- and pH-dependent manner, by prolonging the lag phase without affecting final carrying capacity in vitro. These metabolites further suppressed bacterial swimming motility and biased the orientation of fimS, the phase variable switch for T1 fimbriae, under acidic conditions. In a human polarized, mucus-secreting intestinal infection model, SCFA treatment during UPEC challenge altered bacterial localization patterns, favouring planktonic over mucosal-associated populations, and preserved epithelial barrier function. Together, these in vitro findings demonstrate that SCFAs modulate key UPEC colonization-associated phenotypes and influence host-pathogen dynamics at the colonic epithelial interface. These results provide mechanistic insights into how depletion of SCFA-producing gut commensals may alter the intestinal reservoir environment in vitro and warrants further investigation into the role of gut-derived SCFAs in rUTI susceptibility.

microbiology↗

bioRxiv2023

A human urothelial microtissue model reveals shared colonization and survival strategies between uropathogens and asymptomatic bacteria

Urinary tract infection is among the most common infections worldwide, and is typically studied in animals and cell lines with limited uropathogenic strains. Here, we assessed diverse bacterial pathogens and asymptomatic bacteria (ASB) in a human urothelial microtissue model including full stratification/differentiation and urine tolerance. Several uropathogens and ASB-like E. coli invaded intracellularly, suggesting invasion is a shared survival strategy, instead of a virulence hallmark. The E. coli adhesin FimH was required for intracellular community formation, but not for invasion. Other shared lifestyles included filamentation (Gram-negatives), chaining (Gram-positives) and hijacking of exfoliating cells, while biofilm-like aggregates formed mainly with Pseudomonas and Proteus. Urothelial cells expelled invasive bacteria in Rab-/LC3-decorated structures, while highly cytotoxic/invasive uropathogens, but not ASB, disrupted host barrier function and strongly induced exfoliation and cytokine production. Overall, this work highlights diverse species-/strain-specific infection strategies and corresponding host responses in a human urothelial microenvironment, providing insights at the tissue, cell and molecular level. One-Sentence SummaryA human urothelial model revealed shared colonization strategies between uropathogens and asymptomatic bacteria, and pathogen-specific innate immune responses

microbiology↗

bioRxiv2022

Pathophysiology of Dyt1 dystonia is mediated by spinal cord dysfunction

Dystonia, a neurological disorder defined by abnormal postures and disorganised movements, is considered to be a neural circuit disorder with dysfunction arising within and between multiple brain regions. Given that spinal circuits constitute the final pathway for motor control, we sought to determine their contribution to the movement disorder. Focusing on the most common inherited dystonia, DYT1-TOR1A, we confined a conditional knockout of Tor1a to the spinal cord and dorsal root ganglia (DRG) and found that these mice recapitulated the phenotype of the human condition, developing early onset generalised torsional dystonia. Physiologically, these mice bore the hallmark features of dystonia: spontaneous contractions at rest, excessive sustained contractions during voluntary movements including co-contractions of motor antagonists, and altered sensory-motor reflexes. Furthermore, spinal locomotor circuits were impaired. Together, these data challenge current understanding of dystonia, and lead to broader insights into spinal cord function and movement disorder pathophysiology.

neuroscience↗

bioRxiv2020

Asymmetric nuclear division of neural stem cells contributes to the formation of sibling nuclei with different identities

Cellular diversity in multicellular organisms is often generated via asymmetric divisions. In the fly, for example, neural stem cells divide asymmetrically to generate a large self-renewing stem cell and a smaller sibling that differentiates. Efforts to understand how these different cell fates are generated have focused on the asymmetric segregation of cortically-localised transcription factors at division, which preferentially enter single daughter cell nuclei to change their fate. However, we find that the nuclear compartment in these cells remains intact throughout mitosis and is asymmetrically inherited, giving rise to sibling nuclei that differ profoundly in size, envelope composition and fate markers. These data reveal the importance of considering nuclear remodelling during stem cell divisions, and show how daughter cell fates depend on the coordination of the asymmetric inheritance of cortical fate markers with asymmetric nuclear division.

cell biology↗

bioRxiv2020

Acrylamide Inhibits Vaccinia Virus Through Vimentin-independent Anti-Viral Granule Formation

The replication and assembly of vaccinia virus (VACV), the prototypic poxvirus, occurs exclusively in the cytoplasm of host cells. While the role of cellular cytoskeletal components in these processes remains poorly understood, vimentin - a type III intermediate filament - has been shown to associate with viral replication sites and to be incorporated into mature VACV virions. Here we employed chemical and genetic approaches to further investigate the role of vimentin during the VACV lifecycle. The collapse of vimentin filaments, using acrylamide, was found to inhibit VACV infection at the level of genome replication, intermediate- and late- gene expression. However, we found that CRISPR-mediated knockout of vimentin did not impact VACV replication. Combining these tools, we demonstrate that acrylamide treatment results in the formation of antiviral granules (AVGs) known to mediate translational inhibition of many viruses. We conclude that vimentin is dispensable for poxvirus replication and assembly and that acrylamide, as a potent inducer of AVGs during VACV infection, serves to bolster cells antiviral response to poxvirus infection. Summary StatementAcrylamide inhibits poxvirus replication by inducing anti-viral granules and blocking translation. This inhibition is independent of the effect of acrylamide on vimentin filaments which were found to be dispensable for viral replication and assembly.

microbiology↗

bioRxiv2020

Vaccinia virus hijacks ESCRT-mediated multivesicular body formation for virus egress

Unlike most enveloped viruses, poxvirus egress is a complex process whereby cytoplasmic single membrane-bound virions are wrapped in a cell-derived double membrane. These triple membrane-bound particles, termed intracellular enveloped virions (IEVs), are then released from infected cells by fusion. While the wrapping double membrane is thought to be derived from virus-modified trans-Golgi or early endosomal cisternae, the cellular factors that regulate virus wrapping remain largely undefined. To identify novel cell factors required for this process the prototypic poxvirus, vaccinia virus (VACV), was subjected to a high-throughput RNAi screen directed against cellular membrane trafficking proteins. Focusing on the endosomal sorting complexes required for transport (ESCRT), we demonstrate that ESCRT-III and VPS4 are required for packaging of virus into multivesicular bodies (MVBs). EM-based characterization of these MVB-IEVs showed that they account for half of IEV production indicating that MVBs serve as a second major source of VACV wrapping membrane. These data support a model whereby, in addition to cisternae-based wrapping, VACV hijacks ESCRT-mediated MVB formation to facilitate virus egress and spread.

cell biology↗

bioRxiv2019

Dynamic trafficking and turnover of Jam-C is essential for endothelial cell migration

Junctional complexes between endothelial cells form a dynamic barrier that hinder passive diffusion of blood constituents into interstitial tissues. Re-modelling of junctions is an essential process during leukocyte trafficking, vascular permeability and angiogenesis. However, for many junctional proteins the mechanisms of junctional remodelling have yet to be determined. Here we used receptor mutagenesis, HRP and APEX-2 proximity labelling, alongside light and electron microscopy, to map the intracellular trafficking routes of junctional adhesion molecule-C (Jam-C). We found that Jam-C co-traffics with receptors associated with changes in permeability, such as VE-Cadherin, NRP-1 and 2, but not with junctional proteins associated with the transmigration of leukocytes. Dynamic Jam-C trafficking and degradation is necessary for junctional remodelling during cell migration and angiogenesis. By identifying new trafficking machinery we show that a key point of regulation is the ubiquitylation of Jam-C by the E3 ligase CBL, this controls the rate of recycling versus lysosomal degradation.

cell biology↗

bioRxiv2019

Macrophages enforce the blood nerve barrier

The specialised blood barriers of the nervous system are important for protecting the neural environment but can hinder therapeutic accessibility1,2. Studies in the central nervous system (CNS) have shown the importance of the cellular components of the neuro-vascular unit for blood-brain barrier (BBB) function. Whilst the endothelial cells (ECs) confer barrier function with specialised tight junctions (TJs) and low levels of transcytosis, pericytes and astrocytes provide complete coverage of the ECs and both deliver essential signals for the development and maintenance of the BBB3-9. In contrast, the blood-nerve barrier (BNB) of the peripheral nervous system (PNS) remains poorly defined10. Here, we show that the vascular unit in the PNS has a distinct cellular composition with only partial coverage of the BNB-forming ECs. Using a mouse model, in which barrier function can be controlled11, we show the BNB, while less tight than the BBB, is maintained by low levels of transcytosis and the TJs of the ECs, with opening of the barrier associated with increased transcytosis. Importantly, we find that while ECs of the PNS have higher transcytosis rates than those of the CNS, the barrier is reinforced by resident macrophages that specifically engulf leaked material. This identifies a distinct role for macrophages as an important component of the BNB acting to protect the PNS environment with implications for improving therapeutic delivery to this tissue.

neuroscience↗