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Stimulation of inositol 1,4,5-trisphosphate (IP3) receptor subtypes by analogues of IP3.

ABSTRACT: Most animal cells express mixtures of the three subtypes of inositol 1,4,5-trisphosphate receptor (IP(3)R) encoded by vertebrate genomes. Activation of each subtype by different agonists has not hitherto been examined in cells expressing defined homogenous populations of IP(3)R. Here we measure Ca(2+) release evoked by synthetic analogues of IP(3) using a Ca(2+) indicator within the lumen of the endoplasmic reticulum of permeabilized DT40 cells stably expressing single subtypes of mammalian IP(3)R. Phosphorylation of (1,4,5)IP(3) to (1,3,4,5)IP(4) reduced potency by ~100-fold. Relative to (1,4,5)IP(3), the potencies of IP(3) analogues modified at the 1-position (malachite green (1,4,5)IP(3)), 2-position (2-deoxy(1,4,5)IP(3)) or 3-position (3-deoxy(1,4,5)IP(3), (1,3,4,5)IP(4)) were similar for each IP(3)R subtype. The potency of an analogue, (1,4,6)IP(3), in which the orientations of the 2- and 3-hydroxyl groups were inverted, was also reduced similarly for all three IP(3)R subtypes. Most analogues of IP(3) interact similarly with the three IP(3)R subtypes, but the decrease in potency accompanying removal of the 1-phosphate from (1,4,5)IP(3) was least for IP(3)R3. Addition of a large chromophore (malachite green) to the 1-phosphate of (1,4,5)IP(3) only modestly reduced potency suggesting that similar analogues could be used to measure (1,4,5)IP(3) binding optically. These data provide the first structure-activity analyses of key IP(3) analogues using homogenous populations of each mammalian IP(3)R subtype. They demonstrate broadly similar structure-activity relationships for all mammalian IP(3)R subtypes and establish the potential utility of (1,4,5)IP(3) analogues with chromophores attached to the 1-position.

SUBMITTER: Saleem H

PROVIDER: S-EPMC3556037 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Stimulation of inositol 1,4,5-trisphosphate (IP3) receptor subtypes by analogues of IP3.

Saleem Huma H Tovey Stephen C SC Rahman Taufiq T Riley Andrew M AM Potter Barry V L BV Taylor Colin W CW

PloS one 20130125 1

Most animal cells express mixtures of the three subtypes of inositol 1,4,5-trisphosphate receptor (IP(3)R) encoded by vertebrate genomes. Activation of each subtype by different agonists has not hitherto been examined in cells expressing defined homogenous populations of IP(3)R. Here we measure Ca(2+) release evoked by synthetic analogues of IP(3) using a Ca(2+) indicator within the lumen of the endoplasmic reticulum of permeabilized DT40 cells stably expressing single subtypes of mammalian IP(3 ...[more]

PMID: 23372785

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Project description:The Ins(1,4,5)P3 regioisomers, Ins(1,4,6)P3 and Ins(1,3,6)P3, which can mimic the 1,4,5-arrangement on the inositol ring of Ins(1,4,5)P3, were examined for Ca2+ release by using four types of saponin-permeabilized cell possessing various abundances of receptor subtypes, with special reference to the relation of potency to receptor subtype. Ins(1,4,6)P3 and Ins(1,3,6)P3 were weak agonists in rat basophilic leukaemic cells (RBL cells), which possess predominantly subtype II receptors, with respective potencies of 1/200 and less than 1/500 that of Ins(1,4,5)P3 [the EC50 values were 0.2, 45 and more than 100 microM for Ins(1,4,5)P3, Ins(1,4,6)P3 and Ins(1,3,6)P3 respectively]. Similar rank order potencies were also evaluated for the displacement of [3H]Ins(1,4,5)P3 bound to RBL cell membranes by these regioisomers. However, they caused Ca2+ release from GH3 rat pituitary cells possessing predominantly subtype I receptors more potently; Ins(1,4,6)P3 and Ins(1,3,6)P3 evoked release at respective concentrations of only one-third and one-twentieth that of Ins(1,4,5)P3 (the EC50 values were 0.4, 1.2 and 8 microM for Ins(1,4,5)P3, Ins(1,4,6)P3 and Ins(1,3,6)P3 respectively). In COS-1 African green-monkey kidney cells, with the relative abundances of 37% of the subtype II and of 62% of the subtype III receptor, potencies of 1/40 and approx. 1/200 for Ins(1, 4,6)P3 and Ins(1,3,6)P3 respectively were exhibited relative to Ins(1,4,5)P3 (the EC50 values were 0.4, 15 and approx. 80 microM for Ins(1,4,5)P3, Ins(1,4,6)P3 and Ins(1,3,6)P3 respectively). In HL-60 human leukaemic cells, in spite of the dominant presence of subtype I receptors (71%), similar respective potencies to those seen with COS-1 cells were exhibited (the EC50 values were 0.3, 15 and approx. 100 microM for Ins(1,4,5)P3, Ins(1,4,6)P3 and Ins(1,3,6)P3 respectively). These results indicate that these regioisomers are the first ligands that distinguish between receptor subtypes; the present observations are of significance for the future design of molecules with enhanced selectivity.

Dataset Information

Stimulation of inositol 1,4,5-trisphosphate (IP3) receptor subtypes by analogues of IP3.

Publications

Stimulation of inositol 1,4,5-trisphosphate (IP3) receptor subtypes by analogues of IP3.

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