Subcellular targeting of kinases and phosphatases by association with bifunctional anchoring proteins.

Introduction Since the discovery by Sutherland and colleagues [l-31 of cyclic AMP as an intracellular second messenger for hormone-mediated events, considerable research has focused on understanding its action. A key step in this process was the discovery by Fischer and Krebs that cyclic AMP, M$+ and ATP were factors affecting the activity of phosphorylase b kinase [4], which led to the discovery of a cyclic-AMP-dependent protein kinase (PKA) [ 51. Since then, it has been demonstrated that 20 or more different hormones activate PKA through a common signalling pathway that elevates intracellular cyclic AMP [6-81. PKA is a multifunctional enzyme with a broad substrate specificity. It is therefore hard to imagine how the kinase is able to phosphorylate the correct target proteins rapidly and preferentially in response to activation by individual hormones. This is particularly true if one considers PKA as a soluble enzyme freely diffusing throughout the cytoplasm. Under these conditions, elevation of cyclic AMP concentration would cause an indiscriminate burst of phosphorylation. Thus a preferred hypothesis is that PKA is compartmentalized and individual hormones preferentially activate specific pools of kinase that are co-localized with substrate proteins (Figure 1). Consequently, the localization of PKA may be a key regulatory event in determining the intracellular sites of hormone action. Support for PKA compartmentalization is provided by three lines of evidence: different hormones activate specific PKA subtypes [9], cyclic AMP accumulates in distinct cellular compartments [lo], and PKA subunits are detected in different cellular compartments [ 11,121. Moreover, a body of work over the past 10 years has shown that the type I1 PKA holoenzyme (2R2C) can be tethered at specific subcellular locations through the interaction of its regulatory (RII) subunit with A-kinase anchor proteins (AKAPs) [ 13-16]. The scope of this paper