Activities of protein kinases

Excitation of smooth muscle via alpha-1 receptors (eg, in the utems, vascular smooth muscle) is accompanied by an increase in intraceUular-free calcium, possibly by stimulation of phosphoUpase C which accelerates the breakdown of polyphosphoinositides to form the second messengers inositol triphosphate (IP3) and diacylglycerol (DAG). IP3 releases intracellular calcium, and DAG, by activation of protein kinase C, may also contribute to signal transduction. In addition, it is also thought that alpha-1 adrenergic receptors may be coupled to another second messenger, a pertussis toxin-sensitive G-protein that mediates the translocation of extracellular calcium. 

FIGURE 2.7 Production of second messengers inositol 1,4,5-triphosphate (IP3) and diacylglycerol (DAG) through activation of the enzyme phospholipase C. This enzyme is activated by the a- subunit of Gq-protein and also by Py subunits of Gj-protein. IP3 stimulates the release of Ca2+ from intracellular stores while DAG is a potent activator of protein kinase C. 

Diacylglycerol is glycerol esterified to two fatty acids at the sn-1 and sn-2 positions. It is a membrane-embedded product of phospholipase C action and an activator of protein kinase C. It is also an intermediate in the biosynthesis of triacylglycerol, phosphatidyletha-nolamine and phosphatidylcholine. 

Protein Kinase Inhibitors. Figure 1 Major mechanism by which inhibitors can prevent the activity of protein kinases. 

Just as myosins are able to move along microfilaments, there are motor proteins that move along microtubules. Microtubules, like microfilaments, are polar polymeric assemblies, but unlike actin-myosin interactions, microtubule-based motors exist that move along microtubules in either direction. A constant traffic of vesicles and organelles is visible in cultured cells especially using time-lapse photography. The larger part of this movement takes place on micrombules and is stimulated by phorbol ester (an activator of protein kinase C), and over-expression of N-J aj oncoprotein (Alexandrova et al., 1993). 

DAG is a potent activator of protein kinase C (PKC). It has been long known that ACh causes an increase in the number of effective L-channels in many smooth muscle cells. This effect is mimicked by phorbol esters (known to activate PKC) and DAG itself Therefore, it has been suggested that one of the actions of ACh involves the activation of L-channels via the evoked increase of DAG. 

Chen L, Huang LY Sustained potentiation of NMD A receptor-mediated glutamate responses through activation of protein kinase C by a mu opioid. Neuron 7 319-326, 1991... 

The general picture of muscle contraction in the heart resembles that of skeletal muscle. Cardiac muscle, like skeletal muscle, is striated and uses the actin-myosin-tropomyosin-troponin system described above. Unlike skeletal muscle, cardiac muscle exhibits intrinsic rhyth-micity, and individual myocytes communicate with each other because of its syncytial nature. The T tubular system is more developed in cardiac muscle, whereas the sarcoplasmic reticulum is less extensive and consequently the intracellular supply of Ca for contraction is less. Cardiac muscle thus relies on extracellular Ca for contraction if isolated cardiac muscle is deprived of Ca, it ceases to beat within approximately 1 minute, whereas skeletal muscle can continue to contract without an extraceUular source of Ca +. Cyclic AMP plays a more prominent role in cardiac than in skeletal muscle. It modulates intracellular levels of Ca through the activation of protein kinases these enzymes phosphorylate various transport proteins in the sarcolemma and sarcoplasmic reticulum and also in the troponin-tropomyosin regulatory complex, affecting intracellular levels of Ca or responses to it. There is a rough correlation between the phosphorylation of Tpl and the increased contraction of cardiac muscle induced by catecholamines. This may account for the inotropic effects (increased contractility) of P-adrenergic compounds on the heart. Some differences among skeletal, cardiac, and smooth muscle are summarized in... 

Larsson, R. and Cerutti, P. (1989). Translocation and enhancement of phosphotransferase activity of protein kinase C following exposure of mouse epidermal cells to oxidants. Cancer Res. 49, 5627-5632. 

The effect of stimulation of cardiac adrenoceptors is even more leisurely because several more steps follow activation of the Gs protein by the p-adrenoceptor. For example, to increase the force of cardiac contraction, we have (1) activation of adenylate cyclase by Gas-GTP, (2) formation of cAMP, (3) activation of protein kinase A by the cAMP, then (4) phosphorylation of the calcium channel protein by the kinase. As a result, it takes about 5 to 6 sec from the time the receptors are... 

FIGURE 8.8 Mechanism of activation of protein kinase B (PKB). PI3-kinase is recruited to the membrane via direct association with the receptor PTK or via association with the docking protein Gab-1. It catalyzes the generation of phosphatidyl-3,4,5-inositolphosphate, which serves as a membrane-recruitment signal for PKB. Associated with the membrane, it is first phosphorylated in its catalytic domain by PDK1 and then by PDK2 in the hydrophobic motif. The activated PKB then detaches from the membrane. 

Lead also has been shown to substitute for calcium in the activation of calmodulin, but this requires higher levels of lead than does the activation of protein kinase C. Nevertheless, the affinity of lead for calmodulin is higher than that of calcium. Once activated, calmodulin regulates the activity of certain enzymes and transporters. For example, it activates c-AMP phosphodiesterase to hydrolyze and terminate the action of cAMP, another second messenger (Bressler and Goldstein 1991 Goldstein 1993 Goering 1993). 

The plant is known to contain chelerythrine chloride, which inhibits the aggregation of rabbit platelet in vitro via inhibition on thromboxane formation and phosphoinosi-tides breakdown (30). Chelerythrine, which occurs in members of the family Papaver-aceae, has been reported to inhibit the enzymatic activity of protein kinase C and to exert cell-growth inhibitory effect via the induction of apoptosis in numerous cancer cell lines (31,32). What is the topoisomerase activity of chelerythrine ... 

Chmura SJ, Dolan ME, Cha A, Mauceri HJ, Kufe DW, Weichselbaum RR. In vitro and in vivo activity of protein kinase C inhibitor chelerythrine chlorise induces tumor cell toxicity and growth delay in vivo. Clin Cancer Res 2000 6 737-742. 

The H2 receptor is the second class of HA receptors. This is another G-protein-coupled receptor but, unlike the Hi receptor, the H2 receptor is coupled to adenylyl cyclase via the GTP-binding Gs protein (Hill et ah, 1997). Encoded by an intronless gene and located on human chromosome 5, the H2 receptor is made up of c. 358 amino acids (Gantz et ah, 1991 Traiffort et ah, 1995). Activation of the H2 receptor causes an accumulation of cAMP and activation of protein kinase A that eventually leads to the activation of cyclic-AMP-response element (CRE)-binding protein (CREB) (Hill et ah, 1997). In neurons, the H2 receptor mediates its excitatory effects by blocking the Ca2+-dependent K+ channel (Haas Konnerth, 1983). 

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