Biology subjects

Bun, P.

Publications and source records attributed to Bun, P..

3 recordsLinked to original sources

bioRxiv2022

Versatile and automated workflow for the analysis of oligodendroglial calcium signals in preclinical mouse models of myelin repair

Intracellular Ca2+ signals of oligodendroglia, the myelin-forming cells of the central nervous system, regulate vital cellular processes including myelination. However, studies on oligodendroglia Ca2+ signal dynamics are still scarce, especially during myelin repair, and there are no software solutions to properly analyze the unique Ca2+ signal characteristics in these cells. Here, we provide a comprehensive experimental and analytical workflow to acquire and analyze Ca2+ imaging data of oligodendroglia at the population and single-cell levels in preclinical mouse models of myelin repair. We report diverse ex vivo and in vivo experimental protocols to obtain reproducible Ca2+ imaging data from oligodendroglia in demyelinated lesions. Importantly, we provide an analytical pipeline containing two free, open source and cross-platform software programs, Occam and post-prOccam, that enable the fully automated analysis of one- and two-photon Ca2+ imaging datasets from oligodendroglia obtained by either ex vivo or in vivo Ca2+ imaging techniques. This versatile and accessible experimental and analytical framework, which revealed significant but uncorrelated spontaneous Ca2+ activity in oligodendroglia inside demyelinated lesions, should facilitate the elucidation of Ca2+-mediated mechanisms underlying remyelination and therefore help to accelerate the development of therapeutic strategies for the many myelin-related disorders, such as multiple sclerosis.

neuroscience↗

bioRxiv2022

ExoJ: an ImageJ2/Fiji plugin for automated spatiotemporal detection of exocytosis

Exocytosis is a dynamic physiological process that enables the release of biomolecules to the surrounding environment via the fusion of membrane compartments to the plasma membrane. Understanding its mechanisms is crucial, as defects can compromise essential biological functions. The development of pH-sensitive optical reporters alongside fluorescence microscopy enables the assessment of individual vesicle exocytosis events at the cellular level. Manual annotation represents, however, a time-consuming task, prone to selection biases and human operational errors. Here, we introduce ExoJ, an automated plugin based on ImageJ2/Fiji. ExoJ identifies user-defined genuine populations of exocytic events, recording quantitative features including intensity, apparent size and duration. We designed ExoJ to be fully user-configurable, making it suitable to study distinct forms of vesicle exocytosis regardless of the imaging quality. Our plugin demonstrates its capabilities by showcasing distinct exocytic dynamics among tetraspanins and vesicular SNAREs protein reporters. Assessment of performance on synthetic data demonstrated ExoJ is a robust tool, capable to correctly identify exocytosis events independently of signal-to-noise ratio conditions. We propose ExoJ as a standard solution for future comparative and quantitative studies of exocytosis.

bioinformatics↗

bioRxiv2021

Plasmodium sporozoites require the protein B9 to invade hepatocytes

Plasmodium sporozoites are transmitted to a mammalian host during blood feeding by an infected mosquito and invade hepatocytes for initial replication of the parasite in the liver. This leads to the release of thousands of merozoites into the blood circulation and initiation of the pathogenic blood stages of malaria. Merozoite invasion of erythrocytes has been well characterized at the molecular and structural levels. In sharp contrast, the molecular mechanisms of sporozoite invasion of hepatocytes are poorly characterized. Here we report a new role during sporozoite entry for the B9 protein, a member of the 6-cysteine domain protein family. Using genetic tagging and gene deletion approaches in rodent malaria parasites, we show that B9 is secreted from sporozoite micronemes and is required for productive invasion of hepatocytes. Structural modelling indicates that the N-terminus of B9 forms a beta-propeller domain structurally related to CyRPA, a cysteine-rich protein forming an invasion complex with Rh5 and RIPR in P. falciparum merozoites. We provide evidence that the beta-propeller domain of B9 is essential for protein function during sporozoite entry and interacts with P36 and P52, both also essential for productive invasion of hepatocytes. Our results suggest that, despite using distinct sets of parasite and host entry factors, Plasmodium sporozoites and merozoites may share common structural modules to assemble protein complexes for invasion of host cells.

microbiology↗