title: Direct observation of mobility state transitions in RNA trajectories by sensitive single molecule feedback tracking.
abstract: Observation and tracking of fluorescently labeled molecules and particles in living cells reveals detailed information about intracellular processes on the molecular level. Whereas light microscopic particle observation is usually limited to two-dimensional projections of short trajectory segments, we report here image-based real-time three-dimensional single particle tracking in an active feedback loop with single molecule sensitivity. We tracked particles carrying only 1-3 fluorophores deep inside living tissue with high spatio-temporal resolution. Using this approach, we succeeded to acquire trajectories containing several hundred localizations. We present statistical methods to find significant deviations from random Brownian motion in such trajectories. The analysis allowed us to directly observe transitions in the mobility of ribosomal (r)RNA and Balbiani ring (BR) messenger (m)RNA particles in living Chironomus tentans salivary gland cell nuclei. We found that BR mRNA particles displayed phases of reduced mobility, while rRNA particles showed distinct binding events in and near nucleoli.
authors.affiliation: Institute of Physical and Theoretical Chemistry, Rheinische Friedrich-Wilhelms-University Bonn, 53115 Bonn, Germany spille@uni-bonn.de.
authors.forename: Jan-Hendrik
authors.identifier: http://orcid.org/0000-0001-8493-4721
authors.initials: JH
authors.lastname: Spille
authors.affiliation: Institute of Physical and Theoretical Chemistry, Rheinische Friedrich-Wilhelms-University Bonn, 53115 Bonn, Germany.
authors.forename: Tim P
authors.initials: TP
authors.lastname: Kaminski
authors.affiliation: Institute of Physical and Theoretical Chemistry, Rheinische Friedrich-Wilhelms-University Bonn, 53115 Bonn, Germany.
authors.forename: Katharina
authors.initials: K
authors.lastname: Scherer
authors.affiliation: Institute for Organic Chemistry and Chemical Biology, Buchmann Institute for Molecular Life Sciences, Goethe University Frankfurt, Max-von-Laue-Str. 9, 60438 Frankfurt, Germany.
authors.forename: Jennifer S
authors.initials: JS
authors.lastname: Rinne
authors.affiliation: Institute for Organic Chemistry and Chemical Biology, Buchmann Institute for Molecular Life Sciences, Goethe University Frankfurt, Max-von-Laue-Str. 9, 60438 Frankfurt, Germany.
authors.forename: Alexander
authors.initials: A
authors.lastname: Heckel
authors.affiliation: Institute of Physical and Theoretical Chemistry, Rheinische Friedrich-Wilhelms-University Bonn, 53115 Bonn, Germany spille@uni-bonn.de.
authors.forename: Ulrich
authors.initials: U
authors.lastname: Kubitscheck
mesh_terms: D000465:Algorithms; D000818:Animals; D002683:Chironomidae; D059045:Chromosomal Puffs; D005456:Fluorescent Dyes; D008856:Microscopy, Fluorescence / Q000379:methods*; D009685:Nuclear Envelope / Q000737:chemistry; D017785:Photons; D012313:RNA / Q000032:analysis*; D012333:RNA, Messenger / Q000032:analysis; D012339:RNA, Ribosomal, 28S / Q000032:analysis; D053835:Unilamellar Liposomes / Q000737:chemistry
chemical_list: D005456:Fluorescent Dyes; D012333:RNA, Messenger; D012339:RNA, Ribosomal, 28S; D053835:Unilamellar Liposomes; D012313:RNA