Subcellular localization of specific inositol 1,3,4,5-tetrakis([3H]phosphate) binding sites in rat liver membrane fractions: a comparative evaluation of pH sensitivity and binding characteristics.
Inositol 1,3,4,5-tetrakis([3H]phosphate) ([3H]IP4) binding sites were investigated in plasma membranes, nuclei and microsomes derived from the rat liver. The pH optimum for maximum [3H]IP4 binding was not the same for plasma membranes, pH 7.5, nuclei, pH 6.5, and microsomes, pH 8.0. Evidence is presented demonstrating that inositol 1,3,4,5-tetrakis(phosphate) (IP4) was the most effective inositol phosphate in displacing the binding of the [3H]IP4 in all the membrane fractions studied. Furthermore, the rank order of inhibition in various membrane fractions was identical; i.e., IP5, Ins(3,4,5,6), and IP3. This suggests that similar types of putative IP4 receptor proteins are dealt with in the plasma membranes, nuclei, and microsomes. Scatchard analysis of saturation isotherms revealed a single binding site in the plasma membranes and in the microsomes, whereas two binding sites marked by distinct KD and Bmax values were found in the nuclei. The density of putative IP4 binding sites in the plasma membranes corresponded to that of the high-affinity ones in the nuclei. Microsomes contained fewer binding sites as compared with plasma membranes or nuclei. On the basis of the pH sensitivity of [3H]IP4 binding and the KD and Bmax values in various membrane compartments, it is proposed that inositol 1,3,4,5-tetrakis(phosphate) receptor proteins are similar but not identical in membrane fractions in rat liver. Plasma membrane [3H]IP4 binding was displaced with IP4 and IP6, revealing IC50 values of 8 +/- 2 and 150 +/- 20 nM, respectively, indicating that rat liver plasma membrane IP4 receptor is not clathrin assembly protein AP-2.(ABSTRACT TRUNCATED AT 250 WORDS)
Köppler
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Laboratoire de Neurobiologie des Interactions Cellulaires, Centre de Neurochimie du Centre National de Recherche Scientifique, Strasbourg, France.
Mersel
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Malviya
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D000818:Animals; D001667:Binding, Competitive; D002462:Cell Membrane / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007295:Inositol Phosphates / Q000378:metabolism*; D008099:Liver / Q000378:metabolism* / Q000648:ultrastructure; D008862:Microsomes, Liver / Q000378:metabolism; D008930:Mitochondria, Liver / Q000378:metabolism; D011869:Radioligand Assay; D051381:Rats; D018160:Receptors, Cytoplasmic and Nuclear / Q000032:analysis / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism*
D007295:Inositol Phosphates; D018160:Receptors, Cytoplasmic and Nuclear; C061107:inositol-1,3,4,5-tetrakisphosphate receptor; C081655:inositol-1,3,4,5-tetrakisphosphate