Neuropeptides are essential signaling molecules secreted by dense-core vesicles (DCVs). They contribute to information processing in the brain, controlling a variety of physiological conditions. Defective neuropeptide signaling is implicated in several psychiatric disorders. Here, we provide a protocol for the quantitative analysis of DCV fusion events in rodent neurons using pH-sensitive DCV fusion probes and custom-written analysis algorithms. This method can be used to study DCV fusion mechanisms and is easily adapted to investigate fusion principles of other secretory organelles. For complete details on the use and execution of this protocol, please refer to Persoon et al. (2019).
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http://dx.doi.org/10.1016/j.xpro.2021.100325 | DOI Listing |
Biophys J
December 2024
Cellular and Molecular Physiology, School of Medicine, Yale University, New Haven, Connecticut; Nanobiology Institute, Yale University, West Haven, Connecticut; Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut; Saints-Pères Paris Institute for the Neurosciences (SPPIN), Université de Paris, Centre National de la Recherche Scientifique (CNRS) UMR 8003, Paris, France; Wu Tsai Institute, Yale University, New Haven, Connecticut. Electronic address:
Synaptotagmin-1 (Syt1) is a major calcium sensor for rapid neurotransmitter release in neurons and hormone release in many neuroendocrine cells. It possesses two tandem cytosolic C2 domains that bind calcium, negatively charged phospholipids, and the neuronal SNARE complex. Calcium binding to Syt1 triggers exocytosis, but how this occurs is not well understood.
View Article and Find Full Text PDFbioRxiv
September 2024
Cellular and Molecular Physiology, School of Medicine, Yale University, New Haven, CT.
Synaptotagmin-1 (Syt1) is a major calcium sensor for rapid neurotransmitter release in neurons and hormone release in many neuroendocrine cells. It possesses two tandem cytosolic C2 domains that bind calcium, negatively charged phospholipids, and the neuronal SNARE complex. Calcium binding to Syt1 triggers exocytosis, but how this occurs is not well understood.
View Article and Find Full Text PDFSmall Methods
August 2024
State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Biotechnology, Beijing, 100071, China.
The Toll-like receptor 9 (TLR9) stimulator, CpG oligodeoxynucleotide, has emerged as a potent enhancer of protein subunit vaccines. Incorporating the protein antigen directly with the CpG adjuvant presents a novel strategy to significantly reduce the required dosage of CpG compared to traditional methods that use separate components. In contrast to existing chemical conjugation methods, this study introduces an enzymatic approach for antigen-adjuvant coupling using a recombinant endonuclease DCV fused with SpyTag.
View Article and Find Full Text PDFInvest Ophthalmol Vis Sci
March 2024
Ocular Motility & Vision Neurosciences Laboratory, Cole Eye Institute, Cleveland Clinic, Cleveland, Ohio, United States.
Purpose: Amblyopic and strabismus subjects experience inter-ocular suppression, impaired stereoacuity, and increased fixation instability. The purpose of the study was to investigate factors affecting suppression and stereoacuity and examine their relationship to fixation eye movement (FEM) abnormalities.
Methods: We recruited 14 controls and 46 amblyopic subjects (anisometropic = 18, strabismic = 14, and mixed = 14) and 11 subjects with strabismus without amblyopia.
Chem Sci
November 2023
State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan Provincial Key Laboratory of Biomacromolecular Chemical Biology, Hunan University Changsha 410082 P. R. China
Monitoring the spatiotemporal dynamics of cancer biomarkers within the tumor microenvironment (TME) is critical to understanding their roles in tumorigenesis. Here, we reported a multifunctional fusion protein (collagen-binding domain and duck circovirus tag fused to mCherry, CBD-mCherry-DCV) capable of binding collagen with high affinity and covalently binding specific nucleic acids with exceptional efficiency. We then constructed a chimeric protein-nucleic acid nanodevice (CPNN) using CBD-mCherry-DCV and an aptamer-based sensing module to enable spatially controlled ratiometric imaging of cancer biomarkers in the TME.
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