Authors: Isamu Kameshita, Atsuhiko Ishida and Hitoshi Fujisawa

Source:  FEBS Letters (1999) 456: 249-252

Presenter: Njanoor Narayanan, University of Western Ontario

Presenter’s Summary

Background:

Multifunctional Ca2+/calmodulin-dependent protein kinase (CaMKII) mediates cellular responses induced by increase in second messenger Ca2+ and has been implicated in the control of such essential functions as synaptic transmission, gene transcription, cell growth and contraction of cardiac and smooth muscles. A distinguishing feature of CaMKII is autophosphorylation which converts the enzyme to an autonomous kinase. It has been shown that Ca2+/calmodulin-dependent autophosphorylation of a threonine residue within the regulatory domain (Thr-286) is responsible for the generation of  autonomous activity and that autophosphorylation precedes substrate phosphorylation and is required for full activation of the enzyme. Studies in intact cells indicate that autophosphorylation accompanies physiological activation and autophosphorylation renders the kinase constitutively active. Therefore, dephosphorylation of autophosphorylated CaMKII is of critical importance in the deactivation of CaMKII and reversible regulation of CaMKII-mediated cellular processes. Recently, Ishida et al. (J. Biol.Chem. 273: 1904-1910, 1998) reported the isolation and characterization of a 54 kDa protein phosphatase which dephosphorylates the autophosphorylated CaMKII. This enzyme, designated CaMKPase, exhibited much greater substrate specificity when compared with other multifunctional protein phosphatases (e.g. protein phosphatase 1, 2A and 2C) and required Mn2+ and polycations such as poly-L-lysine for its activity.

Summary of Present Work:

The present paper describes a novel reciprocal regulation of CaMKII and CaMKPase. The authors demonstrate that in the presence of a polycation such as poly-L-lysine, CaMKII phosphorylates CaMKPase and this phosphorylation markedly enhances the ability of CaMKPase to dephosphorylate autophosphorylated CaMKII.  It is also shown that the requirement for a polycation is specific for the phosphorylation of CaMKPase but not other substrates by CaMKII. Apparently, a conformational change in CaMKPase induced by the binding of polycation is required for enzyme activity and renders the phosphorylation site in CaMKPase accessible to CaMKII.
 

Why this paper is important?

Reversibility of cellular regulatory processes is of key importance in the maintenance of normal cell function. In the case of CaMKII, its ability to undergo autophosphorylation and the consequent generation of an autonomous enzyme prolongs CaMKII function beyond the transient elevation of intracellular Ca2+. Mechanisms which may serve to restrain CaMKII from producing persistent activation of its targets are, therefore, of considerable significance. This study has identified a novel “self-restraining” mechanism in which CaMKII-induced phosphorylation of a target substrate, CaMKPase, paves the way for deactivation of CaMKII, in vitro. The findings reported in this paper also demonstrate that operation of this regulatory pathway is dependent on the conformational state of CaMKPase. Major new questions arising from this study are:

1. What are the physiological factors which influence the conformational state of CaMKPase so as to control its accessibility to phosphorylation by CaMKII?

2. Apparently, CaMKII phosphorylates a single site in CaMKPase. Which amino acid is phosphorylated and at which location?

3. Does the reciprocal regulation involving phosphorylation of CaMKPase and dephosphorylation of CaMKII occur in a physiological setting- i.e. in intact cells?

4. Another recent study has characterized a CaMKII inhibitor protein in brain (BH. Chang, S. Mukherji and T. R. Soderling. Proc. Natl. Acad. Sci. USA 95: 10890-10895, 1998). Is there any interaction between the CaMKII inhibitor pathway and CaMKPase-mediated CaMKII deactivation pathway?

Authors’ Abstract

Ca2+/calmodulin-dependent protein kinase phosphatase (CaMKPase) is a protein phosphatase which dephospho-rylates autophosphorylated Ca2+/calmodulin-dependent protein kinase II (CaMKII) and deactivates the enzyme (Ishida, A., Kameshita, I. and Fujisawa, H. (1998) J. Biol. Chem. 273, 1904-1910). In this study, a phosphorylation-dephosphorylation relationship between CaMKII and CaMKPase was examined. CaMKPase was not significantly phosphorylated by CaMKII under the standard phosphorylation conditions but was phosphorylated in the presence of poly-L-lysine, which is a potent activator of CaMKPase. The maximal extent of the phosphorylation was about 1 mol of phosphate per mol of the enzyme and the phosphorylation resulted in an about 2-fold increase in the enzyme activity. Thus, the activity of CaMKPase appears to be regulated through phosphorylation by its target enzyme, CaMKII.