Quantitative Mapping of Endosomal DNA Processing by Single Molecule Counting.

Autor: Prakash V; Department of Chemistry, University of Chicago, Chicago, IL, 60637, USA., Tsekouras K; Department of Physics and School of Molecular Sciences, Arizona State University, Tempe, AZ, 85287, USA., Venkatachalapathy M; Department of Chemistry, University of Chicago, Chicago, IL, 60637, USA., Heinicke L; Single Molecule Analysis Group, Department of Chemistry, University of Michigan, Ann Arbor, MI, 48109-1055, USA., Pressé S; Department of Physics and School of Molecular Sciences, Arizona State University, Tempe, AZ, 85287, USA., Walter NG; Single Molecule Analysis Group, Department of Chemistry, University of Michigan, Ann Arbor, MI, 48109-1055, USA., Krishnan Y; Department of Chemistry, University of Chicago, Chicago, IL, 60637, USA.; Grossman Institute of Neuroscience, Quantitative Biology and Human Behavior, University of Chicago, Chicago, IL, 60637, USA.
Jazyk: angličtina
Zdroj: Angewandte Chemie (International ed. in English) [Angew Chem Int Ed Engl] 2019 Mar 04; Vol. 58 (10), pp. 3073-3076. Date of Electronic Publication: 2019 Feb 06.
DOI: 10.1002/anie.201811746
Abstrakt: Extracellular DNA is engulfed by innate immune cells and digested by endosomal DNase II to generate an immune response. Quantitative information on endosomal stage-specific cargo processing is a critical parameter to predict and model the innate immune response. Biochemical assays quantify endosomal processing but lack organelle-specific information, while fluorescence microscopy has provided the latter without the former. Herein, we report a single molecule counting method based on fluorescence imaging that quantitatively maps endosomal processing of cargo DNA in innate immune cells with organelle-specific resolution. Our studies reveal that endosomal DNA degradation occurs mainly in lysosomes and is negligible in late endosomes. This method can be used to study cargo processing in diverse endocytic pathways and measure stage-specific activity of processing factors in endosomes.
(© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)
Databáze: MEDLINE
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