Live-Cell Fluorescence Imaging Reveals the Dynamics of Protein Kinase CK2 Individual Subunits

Abstract

Protein kinase CK2 is a multifunctional enzyme which has long been described as a stable heterotetrameric complex resulting from the association of two catalytic (α or α′) and two regulatory (β) subunits. To track the spatiotemporal dynamics of CK2 in living cells, we fused its catalytic α and regulatory β subunits with green fluorescent protein (GFP). Both CK2 subunits contain nuclear localization domains that target them independently to the nucleus. Imaging of stable cell lines expressing low levels of GFP-CK2α or GFP-CK2β revealed the existence of CK2 subunit subpopulations exhibiting differential dynamics. Once in the nucleus, they diffuse randomly at different rates. Unlike CK2β, CK2α can shuttle, showing the dynamic nature of the nucleocytoplasmic trafficking of the kinase. When microinjected in the cytoplasm, the isolated CK2 subunits are rapidly translocated into the nucleus, whereas the holoenzyme complex remains in this cell compartment, suggesting an intramolecular masking of the nuclear localization sequences that suppresses nuclear accumulation. However, binding of FGF-2 to the holoenzyme triggers its nuclear translocation. Since the substrate specificity of CK2α is dramatically changed by its association with CK2β, the control of the nucleocytoplasmic distribution of each subunit may represent a unique potential regulatory mechanism for CK2 activity.

ACKNOWLEDGMENTS

We acknowledge J. Lamarre for the critical reading of the manuscript, D. Grunwald for the confocal analysis, and T. Misteli for his helpful comments during the FLIP analysis. We thank M. Yoshida for providing us with LMB and C. Touriol for FGF-2 plasmids.

This work was supported by grants from the INSERM, the CNRS (contract 8BC06G), the Commissariat à l'Energie Atomique, the Association pour la Recherche contre le Cancer (réseau ARECA), and the Ligue Nationale contre le Cancer.

O. Filhol and A. Nueda contributed equally to this publication.