At variance with the great majority of protein kinases that become active only in response to specific stimuli and whose implication in tumors is caused by genetic alterations conferring to them unscheduled activity, the highly pleiotropic Ser/Thr-specific protein kinase CK2 is constitutively active even under normal conditions and no gain-of-function CK2 mutants are known. Nevertheless, CK2 level is abnormally high in cancer cells where it is believed to generate an environment favorable to the development of malignancy, through a mechanism denoted as "non-oncogene addiction." This makes CK2 not only an appealing target to counteract different kinds of tumors but also a valuable marker of cells predisposed to undergo neoplastic transformation owing to the presence in them of CK2 level exceeding a critical threshold. Such a prognostic exploitation of CK2 would imply the availability of methods suitable for the reliable, sensitive, and specific quantification of its activity in biological samples and in living cells. The aim of this chapter is to describe a number of procedures applicable to the quantitative determination of CK2 activity and to provide experimental details designed for rendering these assays as sensitive and selective as possible even in the presence of many other protein kinases. The procedures described roughly fall in three categories: (i) in vitro quantification of CK2 activity in crude biological samples and cell lysates; (ii) in-cell assay of endogenous CK2 activity based on the phosphorylation of reporter substrates; (iii) identification of CK2 targets in malignant and normal cells

Assessment of CK2 Constitutive Activity in Cancer Cells

RUZZENE, MARIA;DI MAIRA, GIOVANNI;TOSONI, KENDRA;PINNA, LORENZO
2010

Abstract

At variance with the great majority of protein kinases that become active only in response to specific stimuli and whose implication in tumors is caused by genetic alterations conferring to them unscheduled activity, the highly pleiotropic Ser/Thr-specific protein kinase CK2 is constitutively active even under normal conditions and no gain-of-function CK2 mutants are known. Nevertheless, CK2 level is abnormally high in cancer cells where it is believed to generate an environment favorable to the development of malignancy, through a mechanism denoted as "non-oncogene addiction." This makes CK2 not only an appealing target to counteract different kinds of tumors but also a valuable marker of cells predisposed to undergo neoplastic transformation owing to the presence in them of CK2 level exceeding a critical threshold. Such a prognostic exploitation of CK2 would imply the availability of methods suitable for the reliable, sensitive, and specific quantification of its activity in biological samples and in living cells. The aim of this chapter is to describe a number of procedures applicable to the quantitative determination of CK2 activity and to provide experimental details designed for rendering these assays as sensitive and selective as possible even in the presence of many other protein kinases. The procedures described roughly fall in three categories: (i) in vitro quantification of CK2 activity in crude biological samples and cell lysates; (ii) in-cell assay of endogenous CK2 activity based on the phosphorylation of reporter substrates; (iii) identification of CK2 targets in malignant and normal cells
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11577/118227
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