title: A pH-independent DNA nanodevice for quantifying chloride transport in organelles of living cells.
abstract: The concentration of chloride ions in the cytoplasm and subcellular organelles of living cells spans a wide range (5-130 mM), and is tightly regulated by intracellular chloride channels or transporters. Chloride-sensitive protein reporters have been used to study the role of these chloride regulators, but they are limited to a small range of chloride concentrations and are pH-sensitive. Here, we show that a DNA nanodevice can precisely measure the activity and location of subcellular chloride channels and transporters in living cells in a pH-independent manner. The DNA nanodevice, called Clensor, is composed of sensing, normalizing and targeting modules, and is designed to localize within organelles along the endolysosomal pathway. It allows fluorescent, ratiometric sensing of chloride ions across the entire physiological regime. We used Clensor to quantitate the resting chloride concentration in the lumen of acidic organelles in Drosophila melanogaster. We showed that lumenal lysosomal chloride, which is implicated in various lysosomal storage diseases, is regulated by the intracellular chloride transporter DmClC-b.
authors.affiliation: National Centre for Biological Sciences, TIFR, GKVK, Bellary Road, Bangalore 560065, India.
authors.forename: Sonali
authors.initials: S
authors.lastname: Saha
authors.affiliation: Department of Chemistry, University of Chicago, 929E, 57th Street, E305A, GCIS, Chicago, Illinois 60637, USA.
authors.forename: Ved
authors.initials: V
authors.lastname: Prakash
authors.affiliation: National Centre for Biological Sciences, TIFR, GKVK, Bellary Road, Bangalore 560065, India.
authors.forename: Saheli
authors.initials: S
authors.lastname: Halder
authors.affiliation: Department of Chemistry, University of Chicago, 929E, 57th Street, E305A, GCIS, Chicago, Illinois 60637, USA.
authors.forename: Kasturi
authors.initials: K
authors.lastname: Chakraborty
authors.affiliation: 1] National Centre for Biological Sciences, TIFR, GKVK, Bellary Road, Bangalore 560065, India [2] Department of Chemistry, University of Chicago, 929E, 57th Street, E305A, GCIS, Chicago, Illinois 60637, USA.
authors.forename: Yamuna
authors.initials: Y
authors.lastname: Krishnan
mesh_terms: D000818:Animals; D015374:Biosensing Techniques / Q000295:instrumentation*; D002478:Cells, Cultured; D002623:Chemistry Techniques, Analytical / Q000295:instrumentation*; D002712:Chlorides / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D004247:DNA / Q000737:chemistry*; D004331:Drosophila melanogaster; D004867:Equipment Design; D019544:Equipment Failure Analysis; D006863:Hydrogen-Ion Concentration; D046228:Microarray Analysis / Q000295:instrumentation; D036103:Nanotechnology / Q000295:instrumentation; D015388:Organelles / Q000378:metabolism*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D002712:Chlorides; D004247:DNA