Novel microscopy-based screening method reveals regulators of contact-dependent intercellular transfer.
Contact-dependent intercellular transfer (codeIT) of cellular constituents can have functional consequences for recipient cells, such as enhanced survival and drug resistance. Pathogenic viruses, prions and bacteria can also utilize this mechanism to spread to adjacent cells and potentially evade immune detection. However, little is known about the molecular mechanism underlying this intercellular transfer process. Here, we present a novel microscopy-based screening method to identify regulators and cargo of codeIT. Single donor cells, carrying fluorescently labelled endocytic organelles or proteins, are co-cultured with excess acceptor cells. CodeIT is quantified by confocal microscopy and image analysis in 3D, preserving spatial information. An siRNA-based screening using this method revealed the involvement of several myosins and small GTPases as codeIT regulators. Our data indicates that cellular protrusions and tubular recycling endosomes are important for codeIT. We automated image acquisition and analysis to facilitate large-scale chemical and genetic screening efforts to identify key regulators of codeIT.
Department of Biomedicine, University of Bergen, Jonas Lies Vei 91, N-5009 Bergen, Norway.
Dominik Michael
DM
Frei
Department of Biomedicine, University of Bergen, Jonas Lies Vei 91, N-5009 Bergen, Norway.
Erlend
E
Hodneland
Department of Biomedicine, University of Bergen, Jonas Lies Vei 91, N-5009 Bergen, Norway.
Ivan
I
Rios-Mondragon
Department of Biomedicine, University of Bergen, Jonas Lies Vei 91, N-5009 Bergen, Norway.
Anne
A
Burtey
Advanced Light Microscopy Facility, European Laboratory of Molecular Biology (EMBL), Meyerhofstraße 1, 69117 Heidelberg, Germany.
Beate
B
Neumann
Advanced Light Microscopy Facility, European Laboratory of Molecular Biology (EMBL), Meyerhofstraße 1, 69117 Heidelberg, Germany.
Jutta
J
Bulkescher
Department of Biomedicine, University of Bergen, Jonas Lies Vei 91, N-5009 Bergen, Norway.
Julia
J
Schölermann
Advanced Light Microscopy Facility, European Laboratory of Molecular Biology (EMBL), Meyerhofstraße 1, 69117 Heidelberg, Germany.
Rainer
R
Pepperkok
Department of Biomedicine, University of Bergen, Jonas Lies Vei 91, N-5009 Bergen, Norway.
Hans-Hermann
HH
Gerdes
Department of Biomedicine, University of Bergen, Jonas Lies Vei 91, N-5009 Bergen, Norway.
Tanja
T
Kögel
D002450:Cell Communication / Q000502:physiology*; D002462:Cell Membrane / Q000502:physiology* / Q000648:ultrastructure; D058948:Cell Tracking / Q000379:methods; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D007365:Intercellular Junctions / Q000502:physiology*; D056656:Lab-On-A-Chip Devices; D018613:Microscopy, Confocal / Q000379:methods*; D057054:Molecular Imaging / Q000379:methods; D009218:Myosins / Q000378:metabolism*
D020558:GTP Phosphohydrolases; D009218:Myosins