Inositol 1,4,5-trisphosphate phospholipase

Figure 43-7. Phospholipase C cleaves PIPj into diacylglycerol and inositol trisphosphate. R, generally is stearate, and Rj is usually arachido-nate. IP3 can be dephosphorylated (to the inactive I-1,4-P2) or phosphorylated (to the potentially active I-1,3,4,5-P4).

Fig. 12. Tentative model of the signal transduction chain that links the perception of pectic fragments to defense responses in carrot cells. Abbreviations apy, heterotrimeric G protein CaM, calmodulin 4CL, 4-coumarate-CoA ligase CTX, cholera toxin FC, fusicoccine GDP-P-S and GTP-y-S, guanosine 5 -0-(2-thiodiphosphate) and guanosine 5 -0-(3-thiotriphosphate) IP3, 1,4,5-inositol trisphosphate PAL, phenylalanine ammonia-lyase PLC, phospholipase C PR, pathogenesis related PTX, pertussis toxin Rc, receptor SP, staurosporine. Activation and inhibition are symbolized by + and -respectively.

The other activity associated with transmembrane receptors is phospholipase C. Phosphatidyl inositol is a membrane phospholipid that after phosphorylation on the head group is found in the membrane as a phos-photidylinostitol bis phosphate. Phospholipase C cleaves this into a membrane associated diacylglycerol (the lipid part) and inositol trisphosphate (IP3, the soluble part). Both play a later role in elevating the level of the second messenger, Ca2+. 

This can be illustrated by known interactions between the cAMP and Ca2+ pathways. A first messenger that initially activates the cAMP pathway would be expected to exert secondary effects on the Ca2+ pathway at many levels via phosphorylation by PKA. First, Ca2+ channels and the inositol trisphosphate (IP3) receptor will be phosphorylated by PKA to modulate intracellular concentrations of Ca2+. Second, phospholipase C (PLC) is a substrate for PKA, and its phosphorylation modulates intracellular calcium concentrations, via the generation of IP3) as well as the activity of PKC, via the generation of DAG, and several types of CAMK. Similarly, the Ca2+ pathway exerts potent effects on the cAMP pathway, for example, by activating or inhibiting the various forms of adenylyl cyclase expressed in mammalian tissues (see Ch. 21). 

The phosphorylated phospholipid, phosphatidylinositol bisphosphate, is present in cell membranes. On hydrolysis by a phospholipase, it produces two products, inositol trisphosphate and diacylglycerol (Figure 11.25), as follows ... 

Figure 12.5 Effector mechanism activation of a membrane-bound phospholipase. An example is activation of a membrane-bound phospholipase which hydrolyses phosphatidylinositol bisphosphate (PIP2) and results in the formation of the two messengers, inositol trisphosphate (IP3) and diacylglycerol (DAG). Messenger IP3 binds to a receptor on the endoplasmic reticulum that results in release of Ca ions into the cytosol. DAG, which remains within the membrane, activates protein kinase-C at the membrane surface. When the kinase leaves the membrane, it is unclear how it remains active or loss of activity is prevented, so that it can phosphorylate proteins in the cytosol or even the nucleus. An example is adrenaline binding to the a-receptor in the liver, in which Ca ions stimulate glycogenolysis.

In Uver, adrenaline binds to the a-receptor, and the hormone-receptor complex activates a membrane-bound phospholipase enzyme which hydrolyses the phospholipid phosphatidylinositol 4,5-bisphosphate. This produces two messengers, inositol trisphosphate (IP3) and diacylglycerol (DAG) (Figure 12.5). The increase in IP3 stimulates release of Ca ions from the endoplasmic reticulum into the cytosol, the effect of which is glycogen breakdown and release into the blood (see Figure 12.5 and Chapter 6). 

Phosphatidylinositol bisphosphate (PIP2) is a component of the cell membrane which is hydrolysed by a phospholipase to produce inositol trisphosphate (IP3). 

Figure 21.6 One mechanism of activation of the cell cycle by a growth factor. Binding of growth factor to its receptor activates membrane-bound phospholipase-C. This hydrolyses phosphati-dylinositol bisphosphate in the membrane to produce the messengers, inositol trisphosphate (IP3) and diacylglycerol (DAG). IP3 results in release of Ca from an intracellular store. The increased Ca + ion concentration activates protein kinases including protein kinase-C (PK-C). DAG remains membrane-bound and also activates protein kinase-C (PK-C) which remains in the activated form as it travels through the cell where it phosphory-lates and activates transcription factors. This results in activation of genes that express enzymes involved in nucleotide synthesis, DNA polymerases and cyclins, which are all reguired for the cell cycle (See Chapter 20 for provision of nucleotides and cyclins for the cell cycle).

Figure 22.13 a-Adrenergic receptor control of contraction of smooth muscle. IP3 represents inositol trisphosphate. Binding of a catecholamine to an a-receptor activates a membrane-bound phospholipase which hydrolyses phosphatidyUnositol bisphosphate within the membrane to produce IP, and diacylglycerol (DAG). IP3 binds a receptor on the sarcoplasmic reticulum in smooth muscle, which activates a Ca ion channel and the cytosolic Ca ion concentration increases, which results in contraction of smooth muscle in arterioles. This results in vasoconstriction and hence decreases blood flow which can leading to an increase in blood pressure. 

The same basic biochemical control mechanism causes contraction of the smooth muscle as well as secretion of aldosterone. The binding of angiotensin to its receptor activates a membrane phospholipase-C. It catalyses the hydrolysis of phosphoinositide phosphatidylinositol bis-phosphate to produce the two intracellular messengers, inositol trisphosphate (IP3) and diacylglycerol (DAG). 

Major effector proteins for G-pro-tein-coupled receptors include adenylate cyclase (ATP intracellular messenger cAMP), phospholipase C (phos-phatidylinositol intracellular messengers inositol trisphosphate and di-acylglycerol), as well as ion channel proteins. Numerous cell functions are regulated by cellular cAMP concentration, because cAMP enhances activity of protein kinase A, which catalyzes the transfer of phosphate groups onto functional proteins. Elevation of cAMP levels inter alia leads to relaxation of smooth muscle tonus and enhanced contractility of cardiac muscle, as well as increased glycogenolysis and lipolysis (p. 

Figure 14-3. Signaling through protein kinase C (PKC). Activated phospholipase C cleaves the inositol phospholipid PIP2 to form both soluble (IP3) and membrane-associated (DAG) second messengers. DAG recruits PKC to the membrane, where binding of calcium ions to PKC fully activates it. To accomplish this, IP3 promotes a transient increase of intracellular concentration by binding to a receptor on the endoplasmic reticulum, which opens a channel allowing release of stored calcium ions. PIP2, phosphatidylinositol 4,5-bisphosphate DAG, diacylglycerol PLC, phospholipase C IP3, inositol trisphosphate.

Most of the known actions of Ang II are mediated by the AT receptor, a Gq protein-coupled receptor. Binding of Ang II to ATi receptors in vascular smooth muscle results in activation of phospholipase C and generation of inositol trisphosphate and diacylglycerol (see Chapter 2). These events, which occur within seconds, result in smooth muscle contraction. 

Three subtypes of vasopressin G protein-coupled receptors have been identified. Via receptors mediate the vasoconstrictor action of vasopressin V , receptors potentiate the release of ACTH by pituitary corticotropes and V 2 receptors mediate the antidiuretic action. Via effects are mediated by activation of phospholipase C, formation of inositol trisphosphate, and increased intracellular calcium concentration. V2 effects are mediated by activation of adenylyl cyclase. 

The signal transduction mechanisms triggered by binding of ET-1 to its vascular receptors include stimulation of phospholipase C, formation of inositol trisphosphate, and release of calcium from the endoplasmic reticulum, which results in vasoconstriction. Conversely, stimulation of PGI2 and nitric oxide synthesis results in decreased intracellular calcium concentration and vasodilation. 

Two pathways from the activated receptor are shown. At the left is activation of phospholipase Cy and formation, at a membrane-bound site, of inositol trisphosphate and diacylglycerol (DAG). The main pathway, in the center, activates Ras with the aid of the G protein Sos. Activated Ras, in turn, activates Raf and successive components of the MAPK cascade. At the right a seven-helix receptor activates both phospholipase C(3 and Ras via interaction with a (3y subunit. (B) A generalized scheme for the MAP kinase pathway. See Seger and Krebs.380... 

Further support for the hypothesis that Ca2+ plays a central role in regulating phytoalexin accumulation is provided by experiments in which the turnover of phosphatidylinositol was measured in the plasma membrane of elicitor-treated carrot cells [17]. The carrot cells were first labelled with [3H]myo-inositol and, after the addition of elicitors, acid extracts of the cells were analyzed chromatographically for the production of inositol trisphosphate (IP3). In cells treated with elicitor, the release of radioactive IP3 increased with time and attained a maximum at 3 - 5 min after treatment. Phospholipase activity responsible for the degradation of phosphorylated phosphatidylinositol increased correspondingly. Several reports have shown that IP3 induces rapid release of Ca2+ from intracellular stores in animal cells [18, 19]. Studies on plant cells have also demonstrated that exogenous IP3 releases Ca2+ from microsomal preparations at micromolar concentrations, although only limited... 

After activation of the TCR, there is induction of Src family tyrosine kinase (p56lek), which phosphorylates phospholipase Oyl. This is followed by the hydrolysis of phosphatidylinositol 4,5-bisphosphate, resulting in the production of diacyl-glycerol (DAG) and inositol trisphosphate (IP3). Protein kinase C is activated by DAG, which phosphorylates Ras. Ras is a GTPase and its phosphorylation induces Raf and initiation of MAP kinase signaling pathway. IP3 is involved in calcium-dependent activation of IL-2 gene expression via nuclear factor of activated T cells (NFAT). 

The large amount of work in this area reported by Russian workers and covered in our previous review [1] has also been reviewed by the Russian workers [284], These glycolipids have recently acquired a vastly increased biological interest since the discovery that various agonists at the cell surface stimulate a phospholipase which releases D-wiyo-inositol 1,4,5-trisphosphate from the membrane-bound phosphatidyl-inositol-4,5-bisphosphate. The released inositol trisphosphate acts as a second messenger by mobilising intra-cellular calcium ions [285-290]. 

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