Biology subjects

Smart, A. J.

Publications and source records attributed to Smart, A. J..

4 recordsLinked to original sources

bioRxiv2024

Analysis of Categorical Data with Logistic Regression and the Cochran-Mantel-Haenszel Tests in Biological Experiments

The choice of statistical test is a fundamentally important one when analyzing experimental data. Here, we consider the question of categorical data, defined by their properties (for example color) rather than by continuous numbering. Using simple and complex example datasets generated from Caenorhabditis elegans research, we conduct a statistical analysis of (1) a rare cellular event involving the formation of a neuronal extrusion called an exopher, and of (2) a variable behavioral response across a timescale. Two tests we use here are the Cochran- Mantel-Haenszel (CMH) test and logistic regression. These two tests pose practical challenges to researchers that include lack of easy access to statistical software and the need for prior programming knowledge. To this end we provide step-by-step tutorials and example code. We emphasize the flexibility of logistic regression in handling both simple and complex datasets, emphasizing the capacity of logistic regression to provide more comprehensive insights into experimental outcomes than simpler tests like CMH. By analyzing real biological examples and demonstrating their analysis with R code, we provide a practical guide for biologists to enhance the rigor and reproducibility of categorical data analysis in experimental studies.

genetics↗

bioRxiv2023

Mechanical force of uterine occupation enables large vesicle extrusion from proteostressed maternal neurons

Large vesicle extrusion from neurons may contribute to spreading pathogenic protein aggregates and promoting inflammatory responses, two mechanisms leading to neurodegenerative disease. Factors that regulate extrusion of large vesicles, such as exophers produced by proteostressed C. elegans touch neurons, are poorly understood. Here we document that mechanical force can significantly potentiate exopher extrusion from proteostressed neurons. Exopher production from the C. elegans ALMR neuron peaks at adult day 2 or 3, coinciding with the C. elegans reproductive peak. Genetic disruption of C. elegans germline, sperm, oocytes, or egg/early embryo production can strongly suppress exopher extrusion from the ALMR neurons during the peak period. Conversely, restoring egg production at the late reproductive phase through mating with males or inducing egg retention via genetic interventions that block egg-laying can strongly increase ALMR exopher production. Overall, genetic interventions that promote ALMR exopher production are associated with expanded uterus lengths and genetic interventions that suppress ALMR exopher production are associated with shorter uterus lengths. In addition to the impact of fertilized eggs, ALMR exopher production can be enhanced by filling the uterus with oocytes, dead eggs, or even fluid, supporting that distention consequences, rather than the presence of fertilized eggs, constitute the exopher-inducing stimulus. We conclude that the mechanical force of uterine occupation potentiates exopher extrusion from proximal proteostressed maternal neurons. Our observations draw attention to the potential importance of mechanical signaling in extracellular vesicle production and in aggregate spreading mechanisms, making a case for enhanced attention to mechanobiology in neurodegenerative disease.

neuroscience↗

bioRxiv2022

Intermediate Filaments Associate with Aggresome-like Structures and Influence Rate of Cellular Expulsion of Neuronal Disease Aggregates

Under conditions of proteostasis disequilibrium, neurons can enhance intracellular and extracellular protective mechanisms to guard against neurotoxicity. In mammals, an intracellular response to severe proteostasis imbalance that results from proteosome inhibition is the formation of juxtanuclear intermediate filament-surrounded, aggregate-filled aggresomes, which sequester threatening aggregates for later disposal via lysosomal degradation. Highly proteo-stressed neurons can also engage the assistance of neighboring cells in aggregate removal by loading threatening materials into large exopher vesicles that are transferred to neighboring cells for remote degradation of contents, a process that has been suggested to be analogous to the process that enables aggregate spreading in the human brain in neurodegenerative disease. In C. elegans these large extruded vesicles are called exophers. Here we document that players involved in aggresome biology are required for the elimination of potentially deleterious materials in neuronal exophers. We show that in proteostressed C. elegans touch receptor neurons, intermediate filament proteins IFD-1 and IFD-2 can assemble into juxtanuclear structures with multiple molecular and cellular characteristics of mammalian aggresomes. IFD-concentrating structures depend upon orthologs of mammalian adapter proteins, dynein motors, and microtubule integrity for aggregate collection into juxtanuclear compartments where they associate with ubiquitinated and neurotoxic polyglutamine expansion proteins. Strikingly, disruption of aggresome-decoration genes encoding IFDs or disruption of the BAG/14-3-3/Hsc70 adapter that promote aggregate loading of aggresome-like organelles, lowers exopher production via a cell autonomous mechanism. Although aggresome-like structures are not mandatory exopher cargo, IFD compartments can be extruded from neurons in exophers, revealing a previously unreported strategy to eliminate neuronal aggresome-like organelles via transfer to neighboring cells. Human IF neurofilament light chain hNFL can partially substitute for C. elegans IFD-2 proteins in promoting exopher production, indicating conservation of the capacity of intermediate filaments to influence neuronal aggregate extrusions across phyla. In sum, we identify a requirement for specific intermediate filaments, counterparts of human biomarkers of neuronal injury and disease and major components of Parkinsons disease Lewy bodies, in C. elegans neuronal aggresome-like organelle formation and large vesicle exopher extrusion from stressed neurons.

cell biology↗

bioRxiv2022

Large vesicle extrusion from C. elegans neurons requires phagocytic interaction via the ARF-6 and CED-1/DRAPER pathways

C. elegans neurons under stress can produce giant vesicles, several microns in diameter, called exophers. Current models suggest that exophers are neuroprotective, providing a mechanism for stressed neurons to eject toxic protein aggregates and organelles. However, little is known of the fate of the exopher once it leaves the neuron. We found that exophers produced by mechanosensory neurons in C. elegans are engulfed by surrounding hypodermal skin cells and are then broken up into numerous smaller vesicles that acquire hypodermal phagosome maturation markers, with vesicular contents gradually degraded by hypodermal lysosomes. Consistent with the hypodermis acting as an exopher phagocyte, we found that exopher removal requires hypodermal actin and Arp2/3, and the hypodermal plasma membrane adjacent to newly formed exophers accumulates dynamic F-actin during budding. Efficient fission of engulfed exopher-phagosomes to produce smaller vesicles and degrade their contents requires phagosome maturation factors SAND-1/Mon1, GTPase RAB-35, the CNT-1 ARF-GAP, and microtubule motor associated GTPase ARL-8, suggesting a close coupling of phagosome fission and phagosome maturation. Lysosome activity was required to degrade exopher contents in the hypodermis but not for exopher-phagosome resolution into smaller vesicles. Importantly, we found that GTPase ARF-6 and effector SEC-10/Exocyst activity in the hypodermis, along with the CED-1 phagocytic receptor, is required for efficient production of exophers by the neuron. Our results indicate that the neuron requires specific interaction with the phagocyte for an efficient exopher response, a mechanistic feature potentially conserved with mammalian exophergenesis, and similar to neuronal pruning by phagocytic glia that influences neurodegenerative disease.

neuroscience↗