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Mapping Absolute DNA Density in Cell Nuclei using Single-molecule Localization Microscopy
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Quantitative Mapping of Endosomal DNA Processing by Single Molecule Counting.

Ved Prakash1, Konstantinos Tsekouras2, Muthukumaran Venkatachalapathy1

  • 1Department of Chemistry, University of Chicago, Chicago, IL, 60637, USA.

Angewandte Chemie (International Ed. in English)
|January 23, 2019
PubMed
Summary
This summary is machine-generated.

Innate immune cells process extracellular DNA using endosomal DNase II. A new single-molecule counting method reveals DNA degradation primarily occurs in lysosomes, not late endosomes.

Keywords:
DNADNase IIlysosomesphotobleachingsingle-molecule counting

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Area of Science:

  • Immunology
  • Cell Biology
  • Molecular Biology

Background:

  • Extracellular DNA triggers innate immune responses.
  • Endosomal DNase II is crucial for DNA degradation and immune signaling.
  • Accurate quantification of DNA processing within specific endosomal compartments is needed.

Purpose of the Study:

  • To develop a method for quantitative, organelle-specific mapping of endosomal DNA processing.
  • To determine the stage-specific localization of DNA degradation within innate immune cells.

Main Methods:

  • A novel single-molecule counting technique utilizing fluorescence imaging.
  • Quantitative analysis of cargo DNA processing within distinct endosomal compartments.

Main Results:

  • Endosomal DNA degradation predominantly occurs in lysosomes.
  • DNA degradation is minimal in late endosomes.
  • The method provides organelle-specific resolution for cargo processing.

Conclusions:

  • The developed method enables precise measurement of endosomal DNA processing.
  • This technique can be applied to various endocytic pathways and processing factors.
  • Understanding DNA degradation sites is key for modeling innate immune responses.