Highlighting protein kinase CK2 movement in living cells.
Protein kinase CK2 has traditionally been described as a stable heterotetrameric complex (alpha2beta2) but new approaches that effectively capture the dynamic behavior of proteins, are bringing a new picture of this complex into focus. To track the spatio-temporal dynamics of CK2 in living cells, we fused its catalytic alpha and regulatory beta subunits with GFP and analog proteins. Beside the mostly nuclear localization of both subunits, and the identification of specific domains on each subunit that triggers their localization, the most significant finding was that the association of both CK2 subunits in a stable tetrameric holoenzyme eliminates their nuclear import (Mol Cell Biol 23: 975-987, 2003). Molecular movements of both subunits in the cytoplasm and in the nucleus were analyzed using different new and updated fluorescence imaging methods such as: fluorescence recovery after photo bleaching (FRAP), fluorescence loss in photo bleaching (FLIP), fluorescence correlation spectroscopy (FCS), and photoactivation using a biphoton microscope. These fluorescence-imaging techniques provide unprecedented ways to visualize and quantify the mobility of each individual CK2 subunit with high spatial and temporal resolution. Visualization of CK2 heterotetrameric complex formation could also be recorded using the fluorescence resonance energy transfer (FRET) technique. FRET imaging revealed that the assembling of this molecular complex can take place both in the cytoplasmic and nuclear compartments. The spatio-temporal organization of individual CK2 subunits and their dynamic behavior remain now to be correlated with the functioning of this kinase in the complex environment of the cell.
Theis-Febvre
Nathalie
N
INSERM EMI 104, Département Réponse et Dynamique Cellulaires, CEA 38054, Grenoble, France.
Martel
Véronique
V
Laudet
Béatrice
B
Souchier
Catherine
C
Grunwald
Didier
D
Cochet
Claude
C
Filhol
Odile
O
D016475:3T3 Cells; D021581:Active Transport, Cell Nucleus; D000818:Animals; D047390:Casein Kinase II / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence; D021122:Protein Subunits / Q000378:metabolism; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D021122:Protein Subunits; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D047390:Casein Kinase II