Title

Protein Kinase CK2 Is a Constitutively Active Enzyme that Promotes Cell Survival: Strategies to Identify CK2 Substrates and Manipulate Its Activity in Mammalian Cells

Document Type

Article

Publication Date

2010

Journal

Methods in Enzymology

Volume

484

First Page

471

Last Page

493

URL with Digital Object Identifier

http://dx.doi.org/10.1016/B978-0-12-381298-8.00023-X

Abstract

Protein kinase CK2 is a constitutively active protein serine/threonine kinase that is ubiquitously expressed and essential for the survival of eukaryotic cells. On the basis of its elevated expression in a number of human cancers and its ability to promote tumorigenesis in transgenic mice, CK2 has emerged as a promising candidate for molecular-targeted therapy. Accordingly, there has been considerable interest in identifying the cellular events that are regulated by CK2 and the cellular substrates of CK2 that are responsible for mediating its actions in cells. Large-scale phosphoproteomics studies are revealing extensive lists of candidate CK2 substrates on the basis that these proteins are phosphorylated at sites conforming to the consensus for phosphorylation by CK2. However, efforts to validate the vast majority of these candidates as bona fide physiological CK2 substrates have been hindered by the lack of systematic strategies to identify its direct substrates and manipulate its activity in intact cells. To overcome these limitations, we describe experimental procedures for isolating CK2 from bacteria and from mammalian cells to enable in vitro phosphorylation of candidate substrates. We also outline strategies for manipulating the levels and activity of CK2 in intact cells. Collectively, the methods that are presented in this chapter should enable the identification and characterization of CK2 substrates and CK2-regulated processes both in vitro and in living cells.

Notes

Grant Support: Canadian Institutes of Health Research