In the inactive state, PKA consists of a complex of two catalytic subunits and two regulatory subunits. The binding of cAMP to the regulatory subunits alters their conformation, causing them to dissociate from the complex. The released catalytic subunits are thereby activated to phosphorylate specific target proteins (Figure 15–26). The regulatory subunits of PKA (also called A-kinase) are important for localizing the kinase inside the cell: special A-kinase anchoring proteins (AKAPs) bind both to the regulatory subunits and to a component of the cytoskeleton or a membrane of an organelle, thereby tethering the enzyme complex to a particular subcellular compartment. Some AKAPs also bind other signaling proteins, forming a signaling complex. An AKAP located around the nucleus of heart muscle cells, for example, binds both PKA and a phosphodiesterase that hydrolyzes cAMP. In unstimulated cells, the phosphodiesterase keeps the local cAMP concentration low, so that the bound PKA is inactive; in stimulated cells, cAMP concentration rapidly rises, overwhelming the phosphodiesterase and activating the PKA. Among the target proteins that PKA phosphorylates and activates in these cells is the adjacent phosphodiesterase, which rapidly lowers the cAMP concentration again. This negative feedback arrangement converts what might otherwise be a prolonged PKA response into a brief, local pulse of PKA activity.
Whereas some responses mediated by cAMP occur within seconds (see Figure 15–24), others depend on changes in the transcription of specific genes and take hours to develop fully. In cells that secrete the peptide hormone somatostatin, regulatory inactive complex of
Figure 15–26 The activation of cyclicAMP-dependent protein kinase (PKA).
The binding of cAMP to the regulatory subunits of the PKA tetramer induces a conformational change, causing these subunits to dissociate from the catalytic subunits, thereby activating the kinase activity of the catalytic subunits. The release of the catalytic subunits requires the binding of more than two cAMP molecules to the regulatory subunits in the tetramer. This requirement greatly sharpens the response of the kinase to changes in cAMP concentration, as discussed earlier (see Figure 15–16). Mammalian cells have at least two types of PKAs: type I is mainly in the cytosol, whereas type II is bound via its regulatory subunits and special anchoring proteins to the plasma membrane, nuclear membrane, mitochondrial outer membrane, and microtubules. In both types, once the catalytic subunits are freed and active, they can migrate into the nucleus (where they can phosphorylate transcription regulatory proteins), while the regulatory subunits remain in the cytoplasm.
signal molecule activated ˜ subunit of stimulatory G plasma protein (G ) membranes activated, phosphorylated CREB
Figure 15–27 How a rise in intracellular cyclic AMP concentration can alter gene transcription. The binding of an extracellular signal molecule to its GPCR activates adenylyl cyclase via Gs and thereby increases cAMP concentration in the cytosol. This rise activates PKA, and the released catalytic subunits of PKA can then enter the nucleus, where they phosphorylate the transcription regulatory protein CREB. Once phosphorylated, CREB recruits the coactivator CBP, which stimulates gene transcription. In some cases, at least, the inactive CREB protein is bound to the cyclic AMP response element (CRE) in DNA before it is phosphorylated (not shown). See Movie 15.2.