Phosphatidylinositol Phospholipase C PI-PLC

Hydrolytic cleavage by phosphatidylinositol phospholipase C (PI-PLC) of the cyclic myo-inositol-l,2-phosphate (cIP) at C-2 is faster than that at C-1. A cyclic monofluorophosphonate has been prepared as a stable analogue of the substrate in 

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A bacterial phosphatidylinositol specific phospholipase C (PI-PLC) had been available for many years before it was demonstrated to strip a number of membrane-bound proteins from eukaryotic cell surfaces [1], Such proteins are anchored by a PI moiety in which the 6 position of inositol is glycosidically linked to glucosamine, which in turn is bonded to a polymannan backbone (Fig. 3-10). The polysaccharide chain is joined to the carboxyl terminal of the anchored protein via amide linkage to ethanolamine phosphate. The presence of a free NH2 group in the glucosamine residue makes the structure labile to nitrous acid. Bacterial PI-PLC hydrolyzes the bond between DAG and phosphati-dylinositols, releasing the water-soluble protein polysac charide-inositol phosphate moiety. These proteins are tethered by glycosylphosphatidylinositol (GPI) anchors. 

The family of heterotrimeric G proteins is involved in transmembrane signaling in the nervous system, with certain exceptions. The exceptions are instances of synaptic transmission mediated via receptors that contain intrinsic enzymatic activity, such as tyrosine kinase or guanylyl cyclase, or via receptors that form ion channels (see Ch. 10). Heterotrimeric G proteins were first identified, named and characterized by Alfred Gilman, Martin Rodbell and others close to 20 years ago. They consist of three distinct subunits, a, (3 and y. These proteins couple the activation of diverse types of plasmalemma receptor to a variety of intracellular processes. In fact, most types of neurotransmitter and peptide hormone receptor, as well as many cytokine and chemokine receptors, fall into a superfamily of structurally related molecules, termed G-protein-coupled receptors. These receptors are named for the role of G proteins in mediating the varied biological effects of the receptors (see Ch. 10). Consequently, numerous effector proteins are influenced by these heterotrimeric G proteins ion channels adenylyl cyclase phosphodiesterase (PDE) phosphoinositide-specific phospholipase C (PI-PLC), which catalyzes the hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP2) and phospholipase A2 (PLA2), which catalyzes the hydrolysis of membrane phospholipids to yield arachidonic acid. In addition, these G proteins have been implicated in... 

When the receptor interacts with its associated G protein, the conformation of the guanine-nucleotide-binding site is altered. The subunits then dissociate, and a phosphatidylinositol-specific phospholipase C (PI-PLC) is activated [5]. The subsequent hydrolysis of phosphatidylinositol bisphosphate then produces inositol triphosphate (IP3) and diacylglycerol (DAG), which are known to be secondary messengers. For example, the water soluble IP3 is released into the cell where its ultimate targets are the calcium storage organelles from which Ca2+ is released [3]. The presence of DAG in cells is known to activate the cellular enzyme protein kinase C (PKC) [6, 7], which phosphorylates a number of cellular... 

Figure 2 Schematic model of the intracellular signaling mechanisms including phosphatidylinositol-specific phospholipase C (PI-PLC) isoforms, inositol-1,4,5-triphosphate (IP3) receptor, and protein kinase C (PKC) in the expression of delta opioid receptor agonist-induced spinal antinociception. In addition, PKC is considered to play a substantial role in an intracellular negative feedback action on the spinal delta opioid receptor-mediated antinociceptive pathway.

R. A. Mortara, L. M. Minelli, F. Vandekerckhove, V. Nussenzwieg, and F. J. Ramalho-Pinto, Phosphatidylinositol-specific phospholipase C (PI-PLC) cleavage of GPI-anchored surface molecules of Trypanosoma cruzi triggers in vitro morphological reorganization of trypomastigotes, J. Eukaryot. Microbiol., 48 (2002) 27-37. 

Bacterial phosphatidylinositol-specific phospholipase C (PI-PLC) (reviewed in ref. [51]) or trypanosomal GPI-specific phospholipase C (GPI-PLC) [52-54] releases GPI-anchored proteins from the cell surface leaving behind diacylglycerol. This cleavage not only solubilizes the protein, but also exposes a phosphoinositol-containing cryptic epitope termed cross-reacting determinant [55]. Reactivity of polyclonal antibodies derived against the cross-reacting determinant (CRD) provide additional evidence for the presence of a GPI anchor. 

Compound 126 has been prepared as an isosteric and isopolar inhibitor of a phosphatidylinositol-specific phospholipase C (PI-PLC) fKsm Bacillus cereus ... 

Synthesis of the first generation C-phosphonate glycosylphos-phatidylinositols (GPI) analogs (427) has been reported by Vishwakarma et al. The key step in this protocol involved coupling of a-pseudodi-saccharide (425) with phosphonic acids (426) in quantitative yield (Scheme 129). It has been also demonstrated that these synthetic probes (427) were resistant to hydrolysis by phosphatidylinositol-specific phospholipase C (PI-PLC) and showed moderate inhibition of the enzyme activity. 

Figure 4 Schematic representation of the Ca2+-transporting systems affecting cellular calcium homeostasis during hormonal stimulation, oq = oq-adrenergic receptor VP = vasopressin receptor PLC = phospholipase C PI = phosphatidylinositol PIP = phospha-tidylinositol-4-phosphate PIP2 = phosphatidylinositol-4,5-biphosphate IP3 = inositol-1,4,5-triphosphate DG = diacylglycerol PKC = protein kinase C. (Modified from Refs. 125 and 285.)...

Fig. 11.13. Insulin receptor signaling. The insulin receptor is a dimer of two membrane-spanning a-(3 pairs. The tyrosine kinase domains are shown in blue, and arrows indicate auto-crossphosphorylation. The activated receptor binds IRS molecules (insulin receptor substrates) and phos-phorylates IRS at multiple sites, thereby forming binding sites for proteins with SH2 domains Grb2, phospholipase C"y(PLC"y), and PI 3-kinase. These proteins are associated with various phosphatidylinositol phosphates (all designated with PIP) in the plasma membrane.

Figure 1.11 Pathways involved in phospholipase C (PLC) cellular signalling. PKA, protein kinase A or cAMP-dependent protein kinases PKC, protein kinase C PI, phosphatidylinositol PIP2, phosphatidylinositol bis-phosphate IP3, inositol triphosphate IP, inositol phosphate DAG, diacylglycerol.

Phosphatidylinositol-4,5-bisphosphate (PtdIns(4,5)P2) pools can be regulated by PI kinases, PtdIns(4,5)P2-phosphatases, phospholipase C (PLC), and lipid transfer and binding proteins. The contribution of each to PtdIns(4,5)P2 pools will depend on the metabolic status of the cell. Enzymes involved in PtdIns(4,5)P2 biosynthesis are shown in Figure 1. Our goal is to convince the reader of the importance of characterizing the metabolic fluxes within the discrete subcellular phospholipid microdomains that make up the lipid signaling pools. 

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