In vivo visualization of actin dynamics and actin interactions by BiFC.
The method of bimolecular fluorescence complementation (BiFC) enables selective visualization of protein interactions. While BiFC complex formation under in vitro conditions is considered to be essentially irreversible, there are hints that under in vivo conditions BiFC complex formation can be reversible. In the present study we used the BiFC method to visualize in vivo actin cytoskeleton dynamics. We demonstrate that in living cells formation of actin/actin BiFC complexes is reversible. Furthermore, we show heterologous binding between actin and protein kinase C delta (PKCdelta). Treatment with phorbol esters caused translocation of actin/PKCdelta complexes from the cytosol to the plasma membrane independent of an intact actin cytoskeleton. Our experiments demonstrate that the BiFC method might be a useful tool to investigate participation of the actin cytoskeleton in regulation of cell function.
Anderie
Ines
I
Department of Physiology, University of the Saarland, Homburg, Saar, Germany. ines.anderie@uniklinikum-saarland.de
Schmid
Andreas
A
D000199:Actins / Q000378:metabolism*; D001426:Bacterial Proteins / Q000378:metabolism; D015153:Blotting, Western; D002273:Carcinogens / Q000494:pharmacology; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D008164:Luminescent Proteins / Q000378:metabolism; D008856:Microscopy, Fluorescence; D010703:Phorbol Esters / Q000494:pharmacology; D051745:Protein Kinase C-delta / Q000378:metabolism*
D000199:Actins; D001426:Bacterial Proteins; D002273:Carcinogens; D008164:Luminescent Proteins; D010703:Phorbol Esters; C054562:yellow fluorescent protein, Bacteria; D051745:Protein Kinase C-delta