Biology subjects

Dickey, B.

Publications and source records attributed to Dickey, B..

3 recordsLinked to original sources

bioRxiv2025

Airway Secretory Cells Contain Both a Perinuclear Golgi Ribbon and Dispersed Golgi Satellites

RationaleFinely tuned production and secretion of the polymeric mucins MUC5AC and MUCB are required for lung health, but knowledge of many details between their translation and their packaging into secretory granules is lacking. ObjectivesTo analyze the structure and function of the Golgi apparatus, a key site of mucin glycosylation, folding, polymerization and packaging, in airway epithelial secretory cells. MethodsLung tissue was obtained from mice stimulated or not with IL-13 to upregulate mucin production, and from normal human lungs. Golgi elements in mouse and human tissue were imaged by high-resolution immunofluorescence microscopy and electron microscopy. Tissue from mice with deletion of both polymeric mucins was also examined. Measurements and Main ResultsBy immunofluorescence microscopy, both mouse and human airway secretory cells contained [~]100 dispersed puncta labeled by markers of medial and trans Golgi cisternae and the trans-Golgi network (TGN), but only a few perinuclear puncta were labeled by markers of cis-Golgi cisternae. By electron microscopy, secretory cells contained both a perinuclear Golgi ribbon and numerous dispersed Golgi stacks, termed satellites. In mucous metaplastic cells, satellites were concentrated among immature mucin granules. Increasing mucin production by cytokine stimulation did not increase the number of TGN puncta, nor did preventing polymeric mucin production by gene deletion reduce TGN puncta. ConclusionsMucin-producing airway secretory cells express an unusual Golgi structure consisting of a conventional perinuclear Golgi ribbon as well as dispersed satellites. While the Golgi satellites are likely an adaptation for mucin production and packaging, their presence is specified developmentally, independent of mucin production.

cell biology↗

bioRxiv2022

Screening of hydrocarbon-stapled peptides for inhibition of calcium-triggered exocytosis

The so-called primary interface between the SNARE complex and synaptotagmin-1 (Syt1) is essential for Ca2+-triggered neurotransmitter release in neuronal synapses. The interacting residues of the primary interface are conserved across different species for synaptotagmins (Syt1, Syt2, Syt9), SNAP-25, and syntaxin-1A homologs involved in fast synchronous release. This Ca2+-independent interface forms prior to Ca2+-triggering and plays a role in synaptic vesicle priming. This primary interface is also conserved in the fusion machinery that is responsible for mucin granule membrane fusion. Ca2+-stimulated mucin secretion is mediated by the SNAREs syntaxin-3, SNAP-23, VAMP8, synaptotagmin-2, and other proteins. Here, we designed and screened a series of hydrocarbon-stapled peptides consisting of SNAP-25 fragments that included some of the key residues involved in the primary interface as observed in high-resolution crystal structures. We selected a subset of four stapled peptides that were highly -helical as assessed by circular dichroism and that inhibited both Ca2+-independent and Ca2+-triggered ensemble lipid-mixing with neuronal SNAREs and Syt1. In a single-vesicle contentmixing assay with reconstituted neuronal SNAREs and synaptotagmin-1 or with reconstituted airway SNAREs and synaptotagmin-2, the selected peptides also suppressed Ca2+-triggered fusion. Taken together, hydrocarbon-stapled peptides that interfere with the primary interface consequently inhibit Ca2+-triggered exocytosis. Our inhibitor screen suggests that these compounds may be useful to combat mucus hypersecretion that is a major cause of airway obstruction in the pathophysiology of COPD, asthma and cystic fibrosis.

pharmacology and toxicology↗

bioRxiv2020

Mucin granules are degraded in the autophagosome-lysosome pathway as a means of resolving airway mucous cell metaplasia

Exacerbations of muco-obstructive airway diseases such as COPD and asthma are associated with epithelial changes termed mucous cell metaplasia (MCM). The molecular pathways triggering MCM have been identified; however, the factors that regulate resolution are less well understood. We hypothesized that the autophagosome-lysosome pathway is required for resolution of MCM by degrading cytoplasmic mucins. We found increased intracellular levels of Muc5ac and Muc5b in autophagy-deficient mice. This difference was not due to defective mucin secretion. Instead, we found that Lamp1-labeled lysosomes surrounded mucin granules of mucous cells indicating that granules were being degraded. Using a model of resolution of mucous cell metaplasia in mice, we found increased lysosomal proteolytic activity that peaked in the days after inflammation. Autophagy-deficient mice had persistent accumulation of mucin granules that failed to decline due to reduced mucin degradation. We applied these findings in vitro to human airway epithelial cells (AECs). Activation of autophagy by mTOR inhibition led to degradation of mucin granules in AECs. Our findings indicate that during peak and resolution phases of MCM, mucin granules can be degraded by autophagy. The addition of mucin degradation to the existing paradigm of production and secretion may more fully explain how the secretory cells handle excess amounts of cytoplasmic mucin and offers a therapeutic target to speed resolution of MCM in airway disease exacerbations. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=142 SRC="FIGDIR/small/130534v2_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@94239org.highwire.dtl.DTLVardef@1dac6dforg.highwire.dtl.DTLVardef@368161org.highwire.dtl.DTLVardef@680b7d_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗