Biology subjects

Nelson, L. T.

Publications and source records attributed to Nelson, L. T..

3 recordsLinked to original sources

bioRxiv2026

Calcium signaling recruits Rab27a and the exocyst complex to promote small extracellular vesicle release in skeletal muscle cells

Extracellular vesicles (EVs) are key mediators of intercellular communication with emerging therapeutic potential. Small EVs (~50-150 nm) released from skeletal muscle (SkM) following exercise improve insulin sensitivity and glucose tolerance in animal models, yet the molecular mechanisms governing their secretion remain poorly understood. Ca2+ signaling has been shown to regulate numerous membrane trafficking events ranging from vesicle transport, membrane repair, and exocytosis of cellular cargo. We investigated how Ca2+ signaling influences the intracellular trafficking and membrane delivery mechanisms necessary for EV release using an in vitro SkM cell model. We found that elevated intracellular Ca2+ stimulates EV secretion through a mechanism requiring activity of the octameric exocyst complex. Using proximity ligation assays, co-immunoprecipitation, structural modeling and proximity-dependent proteomics, we identified dynamic coordination between the exocyst complex and the small GTPase, Rab27a, during Ca2+-stimulated EV release. We found this coordination is modulated by Ca2+ signaling and perturbation of exocyst function. We then used direct binding assays to provide a structural basis for the proximity observed between GTP and GDP-loaded Rab27a and exocyst subunits. Additionally, proximity-dependent proteomics revealed that Ca2+ stimulation promoted the association of specific exocyst subunits with GTP-competent Rab27a, suggesting a nucleotide-state-dependent organization of this trafficking interface in living cells. Finally, we adapted a real-time reporter of EV exocytosis in SkM cells and identified Ca2+-responsive kinases that modulate EV secretion. Collectively, our findings support a model in which Ca2+ signaling reorganizes interactions between Rab27a and the exocyst to promote intracellular trafficking necessary for EV secretion in skeletal muscle cells.

cell biology↗

bioRxiv2026

The exocyst is an insulin-sensitive regulator of amyloid precursor protein trafficking and amyloid-beta generation in neurons

Intracellular trafficking of amyloid precursor protein (APP) critically influences amyloidogenic processing, yet the mechanisms regulating this pathway remain incompletely defined. The exocyst is a highly conserved, insulin-responsive, eight-protein Rab effector complex that directs intracellular transport vesicle targeting and docking. We identified APP in a proteomics screen of neuronal cell surface proteins altered after chemical inhibition of exocyst activity. In SH-SY5Y cells expressing a mutant APP that enhances amyloidogenic processing, RNAi-mediated silencing of exocyst subunits significantly decreased sAPP and A{beta} secretion, leading to significant intracellular APP accumulation. We found high-resolution co-localization of APP with exocyst subunits in soma and neurites of differentiated human SH-SY5Y neurons and mouse primary hippocampal neurons, and live-cell TIRF microscopy identified highly coordinated movement between fluorescently-tagged exocyst and APP proteins. These interactions were confirmed in these cells and in mouse brain histological sections by proximity ligation assays (PLAs) demonstrating close (<40nm) APP-EXOC5 association. To examine if exocyst activity in neurons is regulated by insulin, as it is in adipocytes and muscle, we generated a SH-SY5Y cell line with pHluorin-tagged GLUT4. Inhibition of the exocyst prevented exocytosis of GLUT4 to the plasma membrane in response to insulin. Additionally, using PLAs in mouse primary hippocampal neurons and SH-SY5Y neurons, we found that GLUT4-EXOC5 associations were increased by insulin signaling, but APP-EXOC5 associations were markedly reduced, indicating insulin-dependent retargeting of the exocyst complex away from APP+ vesicles towards GLUT+ vesicles. All together, these data identify the exocyst as a novel insulin-regulated mediator of neuronal APP trafficking and A{beta} secretion. In BriefWe show that the insulin-responsive exocyst regulates amyloidogenic processing of APP in neurons and that insulin signaling shifts the exocyst away from APP trafficking to promote the translocation of GLUT4-containing vesicles to the plasma membrane of neurons.

neuroscience↗

bioRxiv2022

mTOR inhibitor-independent Autophagy Activator Ameliorates Cellular Tauopathy and Prionopathy Neurodegeneration Phenotypes

Autophagy-lysosomal impairment is an early and prominent feature of neurodegeneration. Autophagy activation reduces protein aggregates and lipid level abnormalities. We performed a high-content imaging-based screen assessing 940,000 small molecules to identify those that reduce lipid droplet numbers. Of 77 validated, structurally diverse hits, 24 increased autophagy flux reporter activity, consistent with accelerated lipid droplet clearance by lipophagy. Of these, we show that CCT020312 activates autophagy independently of mammalian target of rapamycin (mTOR) inhibition, to avoid immunosuppression. CCT020312 reduced insoluble phosphorylated tau levels and tau-mediated neuronal stress vulnerability, as well as reducing intracellular A{beta} levels within directly induced neurons bearing epigenetic marks of aging derived from Alzheimers patient fibroblasts. Moreover, CCT020312 cleared mutant prion protein aggregates and normalized trafficking deficiencies in axons of a cellular model of familial prion disease. Autophagy is widely considered a promising strategy to attenuate neurodegeneration, and here we introduce a strategy to discover new pharmacology.

cell biology↗