Cytosolic inositol phosphat

Increase in inositol phosphates and cytosolic [Ca2+] Activation of MAPK pathway NN SR 142948A ... 

Human Stably transfected CHO cells Increase in inositol phosphates, cytosolic [Ca2+], arachidonic acid activation of MAPK pathway SR 48692 NT ... 

These effects of ATP are blocked by pertussis toxin, and so the putative ATP receptor is G-protein linked. ATP addition results in phospholipase C activation, which may be detected as increased inositol phosphate metabolism and subsequent elevations in cytosolic free Ca2+. Purinergic receptors on many types of cells are classified as type Pi or P2. Neutrophils possess P2-type receptors, which are activated by ATP and ADP, and also Pi-type receptors, which are activated by adenosine. Occupancy of P2-type receptors enhances fMet-Leu-Phe-mediated effects, whilst occupancy of Pi-type receptors has the opposing effect. Some pharmacological evidence suggests that the P2-type receptor on neutrophils is distinct from the P2X and P2y subtypes that have been described in other cell types. 

Note that equation 5.2 is irreversible and the product AMP will require two phosphorylation steps to reconstitute the high-energy adenosine triphosphate, ATP. Inositol 1,4,5-triphosphate is an important molecule in the cytosol, where it releases calcium ions from storage. It forms part of a series of inositol-phosphate species that mediate calcium ion concentrations inside and outside the cell. 

Selected entries from Methods in Enzymology [vol, page(s)] Chelation, 238, 74, 76, 297 buffers [for analysis of exocytosis, 221, 132 preparation, 219, 186 modulation of cytosolic buffering capacity with quin2, 221, 159] fluorescence assay, 240, 724-725, 740-742 fluorescence imaging, 225, 531 238, 303-304, 322-325, 334-335 free intracellular levels after bacterial invasion, 236, 482-489 free calcium in solutions for membrane fusion analysis, calculation and control, 221, 149 homeostasis mechanisms, 238, 80 hormonal elevation, 238, 79 inositol phosphate effect on release, 238, 207 determination of cytosolic levels [computer methods, 238, 73-75 with fura-2, 238, 73, 146 with indo-1, 238, 298, 316-317 with quin-2, 238, 297] hormone effects, 238, 79 ionomycin effects, 238, 79 membrane depolarization effects,... 

A role as a recognition surface in connecting proteins, similar to the role of SH2 and SH3 domains, has been postulated for PH domains. However, protein partners specifically recognizing PH domains are rare. Therefore, a role of PH domains in anchoring soluble proteins to membranes succeeded the original assumption that PH domains determine protein-protein interactions. In that case, PH domains would be like covalently attached fatty-acid residues, which also help to attach proteins to the membrane. The structural properties of PH domains support a role in bringing cytosolic proteins to the membrane. The structure of a PH domain is shown in Plate 4a. Structures of complexes of inositol phosphates with a PH domain were solved, both by X-ray crystallography and The structural feamres are shown in Plate 4b. 

The most important second messengers are - hydrophilic, cytosolic cAMP, cGMP inositol phosphates... 

Gomisin C has an inhibitory effect on the respiratory burst of rat neutrophils in vitro [243], The mechanism of action may be mediated partly by the suppression of NADPH oxidase and partly by the decrease in cytosolic Ca2+ released from an agonist-sensitive intracellular store. In fact, gomisin C attenuated the activity of TPA-activated neutrophil particulate NADPH oxidase in a concentration-dependent manner and reduced the increase in cytosolic free Ca2+ in neutrophils stimulated by fMLP in presence or absence of ethylenediaminetetraacetic acid (EDTA). In addition, this study suggests that the gomisin C mechanism is not mediated by changes in cellular cAMP or in inositol phosphates, or by scavenging... 

Figure 1. Simplified schematic of receptor-mediated signal transduction in neutrophils. Binding of ligand to the receptor activates a guanine-nucleotide-binding protein (G protein), which then stimulates phospholipase C. Phosphatidylinositol 4,5-bis-phosphate is cleaved to produce diacylglycerol (DAG) and inositol 1,4,5-trisphosphate (IP3). DAG stimulates protein kinase C. IP3 causes the release of Ca from intracellular stores, which results in an increase in the cytosolic Ca concentration. This increase in Ca may stimulate protein kinase C, calmodulin-dependent protein kinases, and phospholipase A2. Protein phosphorylation events are thought to be important in stimulating degranulation and oxidant production. In addition, ionic fluxes occur across the plasma membrane. It is possible that phospholipase A2 and ionic channels may be governed by G protein interactions. ...

The binding of the sperm to a receptor on the membrane of the oocyte either activates a membrane-bound phospholipase or releases a phospholipase into the oocyte. The phospholipase hydrolyses phosphatidylinositol bisphos-phate to produce the two intracellular signals, inositol tris-phosphate (IP3) and diacylglycerol within the ovum. As in other cells, the IP3 signal increases the level of cytosolic Ca + ions and the diacylglycerol (DAG) signal activates protein kinase C. 

Phospholipase C hydrolyzes the bond between glycerol and phosphate in phosphatidylinositol 4,5-bisphos-phate, releasing two products inositol 1,4,5-trisphos-phate (IP3), which is water-soluble, and diacylglycerol, which remains associated with the plasma membrane. IP3 triggers release of Ca2+ from the endoplasmic reticulum, and the combination of diacylglycerol and elevated cytosolic Ca2+ activates the enzyme protein kinase C. 

Abbreviations [Ca2+]j, intracellular Ca2+ concentration as measured by a Ca2 indicator such as ae-quorin [Ca2+]c, Ca2+ concentration in the bulk cytosol (hypothetical value) [Ca2 f]sm, Ca2 concentration in submembrane domain just beneath the plasma membrane (a hypothetical value) PI, phospha-tidylinositol PIP2, phosphatidylinositol 4,5-bisphosphate PIP, phosphatidylinositol 4-phosphate Insl,4,5,P3, inositol 1,4,5,-trisphosphate Ins 1,3,4,P, inositol 1,3,4-trisphosphate Insl,3,4,5P4, inositol 1,3,4,5-tetrakisphosphate Insl,4P2, inositol 1,4-bisphosphate CaM, calmodulin C-kinase, protein kinase C [cAMP]c, cAMP concentration in the bulk cytosol [cAMP]sm, cAMP concentration in submembrane domain just beneath the plasma membrane. 

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