Cyclic AMP-dependent protein kinase (PKA or cAK) and cyclic GMP-dependent protein kinase (PKG or cGK) transfer the γ-phosphate of ATP to serine and threonine residues of many cellular proteins. PKAs are present in most cells and function as effectors of many cAMP-elevating first messengers such as hormones and neurotransmitters. PKG is highly expressed in special cell types such as smooth muscle cells, platelets and cerebellar Purkinje cells (PKG I), as well as intestinal, kidney and brain cells (PKG II). cGMP-elevating agents include NO, natriuretic peptides and guanylin. In most tissues, PKGs are much less abundantly expressed than PKAs.
In the absence of its activating ligand cAMP, PKA exists as an inactive holoenzyme of two regulatory (R) and two catalytic (C) subunits. Following an increase in intracellular cAMP, the R-subunits bind cAMP resulting in the dissociation of the holoenzyme and the release of two free active C-subunits. Active C-subunit phosphorylates peptide substrates containing the -R-R/K-X-S/T- substrate consensus amino acid sequence (although exceptions to this consensus sequence have been observed). The PKA type I holoenzymes (RIα2C2, RIβ2C2) are predominantly cytoplasmic, whereas the majority of type II PKA (RIIα2C2, RIIβ2C2) associates with cytoskeletal structures, organelles and membranes. The holoenzymes can be anchored to specific compartments via interaction of their regulatory subunits with specific PKA anchoring proteins (AKAPs - most of which are identified for PKA II so far).
In contrast to PKA, the regulatory and catalytic regions of the PKG enzyme are present in one polypeptide. Binding of cGMP to the two cGMP-binding sites is thought to release the autoinhibitory N-terminal domain from binding to the C-terminal catalytic domain, thus enabling substrate binding and heterophosphorylation. The substrate consensus amino acid sequence for PKGs appears to require more multiple basic residues than does PKA (consensus -R/K2-3-X-S/T-). However, in vitro, many substrate proteins can be phosphorylated by both kinases. In addition to phosphorylating other proteins (heterophosphorylation), each of the PKGs and type II PKA phosphorylate themselves (autophosphorylation). Within the cell, the specific localization of the kinases and their substrates has been shown to restrict some of the possible interactions suggested by in vitro data. PKG I is localized mainly in the cytoplasm and a number of PKG anchoring proteins (GKAPs, especially for the type Iα enzyme) have been identified. The PKG II enzyme is anchored to membranes via its myristoylated N-terminus.
PKA has been shown to mediate many cellular responses to the intracellular second messenger cAMP in eukaryotes. Other important effectors of cAMP are the guanine nucleotide exchange factors Epac 1 and 2, which activate Rap1, a member of the Ras family of small GTPases. cAMP is also known to directly regulate ion channels. PKA I has been shown to mediate cAMP effects on inhibition of lymphocyte cell proliferation and immune response as well as long term depression in the hippocampus and sensory nerve transmission. PKA II is involved in the regulation of neuronal gene expression and motor learning as well as lipolysis and sperm motility. However, differences between PKA I and II functions are probably mainly due to differences in levels of expression in specific cells and the relative ability of each kinase subtype to localize near substrates by binding to scaffolding proteins (AKAPs). For example PKA II can be localized via AKAPs to the Golgi-centrosomal area, to receptors and ion channels, to the cytoskeleton and the nucleus.
The second messenger cGMP has three major effector systems within the cell: cGMP-regulated ion channels, cGMP-regulated phosphodiesterases and PKGs. PKG I mediates cGMP-induced smooth muscle cell relaxation and inhibition of platelet aggregation. These effects correlate at least partly with an inhibition of calcium release from intracellular stores. In addition PKG I can inhibit cardiac myocyte contractility and has also been shown to regulate proliferation and gene expression in various cell types. PKG II stimulates intestinal chloride secretion, inhibits renin release from juxtaglomerular cells, stimulates renal calcium reabsorption and regulates endochondral ossification.
a) cAMP and most cAMP analogs also activate Epac. To distinguish PKA and Epac effects in intact cells the Epac-specific activators 8-pCPT-2’-O-Me-cAMP or 8-Br-2’-O-Me-cAMP can be used.
b) 6-modified cAMP analogs have a higher affinity for PKA over Epac.
AHA: Aminohexyl amino
AKAP: PKA-anchoring protein
BKCa: Ca2+-regulated potassium channel
6-Bnz-cAMP: N6-Benzoyladenosine-3’,5’-cyclic monophosphate
cAMP: Adenosine-3’, 5’-cyclic monophosphate
cAMPS: Adenosine-3’, 5’-cyclic monophosphorothioate
CFTR: Cystic fibrosis transmembrane conductance regulator
cGMP: Guanosine-3’, 5’-cyclic monophosphate
cGMPS: Guanosine-3’, 5’-cyclic monophosphorothioate
Epac: Exchange protein directly activated by cAMP
GKAP: PKG-anchoring protein
IRAG: IP3-receptor-associated cGMP kinase substrate
KT5720: (9S,10R,12R)-2,3,9,10,11,12-Hexahydro-10-hydroxy-9-methyl-1-oxo-9,12-epoxy-1H-diindolo[1,2,3-fg:3’,2’,1’-kl]pyrrolo[3,4-i][1,6]benzodiazocine-10-carboxylic acid hexyl ester
KT5823: (9S,10R,12R)-2,3,9,10,11,12-Hexahydro-10-methoxy-2,9-dimethyl-1-oxo-9,12-epoxy-1H-diindolo[1,2,3-fg:3’,2’,1’-kl]pyrrolo[3,4-i][1,6]benzodiazocine-10-carboxylic acid methyl ester
8-MA-cAMP: 8-Methylaminoadenosine-3’,5’-cyclic monophosphate
MAP: Microtubule-associated protein
MBS: Myosin-binding subunit of myosin light chain phosphatase
NHERF2: Na+/H+ exchanger regulatory factor 2
PET-cGMP: β-Phenyl-1,N2-ethenoguanosine 3’,5’-monophosphate
8-PIP-cAMP: 8-Piperinoadenosine 3’,5’-cyclic monophosphate
PKA: cAMP-dependent protein kinase
PKG: cGMP-dependent protein kinase
PKI: cAMP-dependent protein kinase inhibitor peptide
RGS-2: Regulator of G-protein signaling-2
Rp-8-Br-cAMPS: 8-Bromoadenosine-3’,5’-cyclic monophosphorothioate, Rp isomer
Rp-8-Cl-cAMPS: 8-Chloroadenosine-3’,5’-cyclic monophosphorothioate, Rp isomer
Rp-cAMPS: Adenosine 3’,5’-cyclic monophosphothioate, Rp-isomer
Rp-8-pCPT-cAMPS: 8-(4-Chlorophenylthio)adenosine-3’,5’-cyclic monophosphorothioate, Rp isomer
Rp-8-Br-PET-cGMPS: b-Phenyl-1,N2-etheno-8-bromoguanosine-3’,5’-cyclic monophosphorothioate, Rp isomer
Rp-8-pCPT-cGMPS: 8-(4-Chlorophenylthio)guanosine-3’,5’-cyclic monophosphorothioate, Rp isomer
Sp-5,6-DCl-cBIMPS: 5, 6-Dichloro-1-b-D-ribofuranosylbenzimidazole-3’, 5’-cyclic monophosphorothioate, Sp-isomer
Sp-8-pCPT-cAMPS: 8-(4-Chlorophenylthio)adenosine-3’,5’-cyclic monophosphorothioate, Sp-isomer
STa: heat-stable enterotoxin secreted by enteropathogenic bacteria
VASP: vasodilator-stimulated phosphoprotein
WASP: Wiskott-Aldrich syndrome protein