Role of Plant-Specific N-Terminal Domain of Maize CK2β1 Subunit in CK2β Functions and Holoenzyme Regulation

Protein kinase CK2 is a highly pleiotropic Ser/Thr kinase ubiquituous in eukaryotic organisms. CK2 is organized as a heterotetrameric enzyme composed of two types of subunits: the catalytic (CK2 alpha) and the regulatory (CK2 beta). The CK2 beta subunits enhance the stability, activity and specificity of the holoenzyme, but they can also perform functions independently of the CK2 tetramer. CK2 beta regulatory subunits in plants differ from their animal or yeast counterparts, since they present an additional specific N-terminal extension of about 90 aminoacids that shares no homology with any previously characterized functional domain. Sequence analysis of the N-terminal domain of land plant CK2 beta subunit sequences reveals its arrangement through short, conserved motifs, some of them including CK2 autophosphorylation sites. By using maize CK2 beta 1 and a deleted version (Delta NCK2 beta 1) lacking the N-terminal domain, we have demonstrated that CK2 beta 1 is autophosphorylated within the N-terminal domain. Moreover, the holoenzyme composed with CK2 alpha 1/Delta NCK2 beta 1 is able to phosphorylate different substrates more efficiently than CK2 alpha 1/CK2 beta 1 or CK2 alpha alone. Transient overexpression of CK2 beta 1 and Delta NCK2 beta 1 fused to GFP in different plant systems show that the presence of N-terminal domain enhances aggregation in nuclear speckles and stabilizes the protein against proteasome degradation. Finally, bimolecular fluorescence complementation (BiFC) assays show the nuclear and cytoplasmic location of the plant CK2 holoenzyme, in contrast to the individual CK2 alpha/beta subunits mainly observed in the nucleus. All together, our results support the hypothesis that the plant-specific N-terminal domain of CK2 beta subunits is involved in the down-regulation of the CK2 holoenzyme activity and in the stabilization of CK2 beta 1 protein. In summary, the whole amount of data shown in this work suggests that this domain was acquired by plants for regulatory purposes.