Biology subjects

Allan, V. J.

Publications and source records attributed to Allan, V. J..

5 recordsLinked to original sources

bioRxiv2025

Auxin-inducible degradation of UNC-116 in C. elegans inhibits bidirectional dense core vesicle transport and worm locomotion on different timescales

The microtubule motor kinesin-1 is vital in neurons, with mutations being associated with neurological diseases. Deletion of the Caenorhabditis elegans kinesin-1 gene unc-116 is lethal, and viable mutants are uncoordinated. Here, we use auxin-mediated degradation to deplete UNC-116 protein at different developmental stages and monitor the effects on cargo transport and locomotion. UNC-116 is substantially degraded within 1 hour of auxin treatment, by which time bidirectional dense core vesicle (DCV) motility is affected. After 4 hours, dynein-driven DCV movement is lost and only limited plus-end-directed DCV motility remains, likely driven by kinesin-3 (UNC-104). DCV movement recovers substantially after rescue from auxin for 24 hours. There is a time-lag between loss of protein and effects on locomotion, as crawling and swimming/thrashing is unaffected until 6-14 hours on auxin. By 18-24 hours, animals are as uncoordinated as the unc-116(rh24sb79) mutant. Notably, degradation of UNC-116 in neurons alone inhibits crawling and swimming, revealing the importance of neuronal kinesin-1 for locomotion. Overall, by bypassing early developmental UNC-116 functions, we reveal that UNC-116 is essential for bidirectional DCV transport and crucial for locomotion.

cell biology↗

bioRxiv2024

The role of Kinesin-1 in neuronal dense core vesicle transport and lifespan regulation in C. elegans

Fast axonal transport is crucial for neuronal function and is driven by kinesins and cytoplasmic dynein. We investigated the role of the kinesin-1 motor complex in dense core vesicle (DCV) transport in C. elegans, using mutants in kinesin light chains (klc-1 and klc-2) and the kinesin motor subunit (unc-116) expressing an ida-1::gfp transgene that labels DCVs in the ALA neuron. A reduced-function unc-116(rf) mutation greatly impaired DCV transport in both directions. A klc-2(rf) reduced-function mutation decreased DCV velocity in both directions and reduced the frequency of body bends during swimming. In contrast, the klc-1(-) null mutation had no effect on anterograde transport or swimming ability, but surprisingly it increased the speed of retrograde DCV transport. We also determined lifespan, finding that klc-1(-) or klc-2(rf) single mutants were wild-type whereas the unc-116(rf), ida-1::gfp and unc-116(rf); ida-1::gfp strains were short-lived. Strikingly, the ida-1::gfp transgenic synergistically interact with either klc mutant to extend lifespan compared to wild-type and parental strains. Our findings suggest that kinesin-1 not only influences anterograde and retrograde DCV transport but also plays a role in regulating lifespan.

cell biology↗

bioRxiv2022

The meiotic LINC complex component KASH5 is an activating adaptor for cytoplasmic dynein

Cytoplasmic dynein-driven movement of chromosomes during prophase I of mammalian meiosis is essential for synapsis and genetic exchange. Dynein connects to chromosome telomeres via KASH5 and SUN1 or SUN2, which together span the nuclear envelope. Here, we show that KASH5 promotes dynein motility in vitro, and cytosolic KASH5 inhibits dyneins interphase functions. KASH5 interacts with either dynein light intermediate chain (DYNC1LI1 or DYNC1LI2) via a conserved helix in the LIC C-terminal, and this region is also needed for dyneins recruitment to other cellular membranes. KASH5s N-terminal EF-hands are essential, as the interaction with dynein is disrupted by mutation of key calcium-binding residues, although it is not regulated by cellular calcium levels. Dynein can be recruited to KASH5 at the nuclear envelope independently of dynactin, while LIS1 is essential for dynactin incorporation into the KASH5-dynein complex. Altogether, we show that the trans-membrane protein KASH5 is an activating adaptor for dynein, and shed light on the hierarchy of assembly of KASH5-dynein-dynactin complexes.

cell biology↗

bioRxiv2020

Network Organisation and the Dynamics of Tubules in the Endoplasmic Reticulum

The endoplasmic reticulum (ER) is a eukaryotic subcellular organelle composed of tubules and sheet-like areas of membrane connected at junctions. The tubule network is highly dynamic and undergoes rapid and continual rearrangement. There are currently few tools to evaluate network organisation and dynamics. We quantified ER network organisation in Vero and MRC5 cells, and developed a classification system for ER dynamics in live cells. The persistence length, tubule length, junction coordination number and angles of the network were quantified. Hallmarks of imbalances in ER tension, indications of interactions with microtubules and other subcellular organelles, and active reorganisation and dynamics were observed. Live cell ER tubule dynamics were classified using a Gaussian mixture model, defining tubule motion as active or thermal and conformational phase space analysis allowed this classification to be refined by tubule curvature states. STATEMENT OF SIGNIFICANCEThe endoplasmic reticulum (ER), a subcellular organelle, is an underexplored real-world example of active matter. Many processes essential to cell survival are performed by the ER, the efficacy of which may depend on its organisation and dynamics. Abnormal ER morphology is linked to diseases such as hereditary spastic paraplegias and it is possible that the dynamics are also implicated. Therefore, analysing the ER network in normal cells is important for the understanding of disease-related alterations. In this work, we outline the first thorough quantification methods for determining ER organisation and dynamics, deducing that tubule motion has a binary classification as active or thermal. Active reorganisation and dynamics along with indications of tension imbalances and membrane contact sites were observed.

biophysics↗

bioRxiv2019

Deciphering anomalous heterogeneous intracellular transport with neural networks

Biological intracellular transport is predominantly heterogeneous in both time and space, exhibiting varying non-Brownian behaviour. Characterisation of this movement through averaging methods over an ensemble of trajectories or over the course of a single trajectory often fails to capture this heterogeneity adequately. Here, we have developed a deep learning feedforward neural network trained on fractional Brownian motion, which provides a novel, accurate and efficient characterization method for resolving heterogeneous behaviour of intracellular transport both in space and time. Importantly, the neural network requires significantly fewer data points compared to established methods, such as mean square displacements, rescaled range analysis and sequential range analysis. This enables robust estimation of Hurst exponents for very short time series data, making possible direct, dynamic segmentation and analysis of experimental tracks of rapidly moving cellular structures such as endosomes and lysosomes. By using this analysis, we were able to interpret anomalous intracellular dynamics as fractional Brownian motion with a stochastic Hurst exponent.

cell biology↗