Perturbation of the human T-cell antigen receptor-T3 complex leads to the production of inositol tetrakisphosphate: evidence for conversion from inositol trisphosphate.

Abstract

Antibodies directed against the T-cell antigen receptor-T3 complex mimic antigen and lead to cellular changes consistent with activation. When cells of the human T-cell line Jurkat were stimulated with a monoclonal antibody directed against T3, inositol phosphates were produced. In addition to inositol trisphosphate, which is the product of phosphatidylinositol bisphosphate cleavage, a second inositol polyphosphate was formed. This compound was more polar than inositol trisphosphate but less polar than inositol pentakisphosphate. It cochromatographed with inositol tetrakisphosphate from ostrich erythrocytes. In permeabilized Jurkat cells, this compound was shown to be formed from inositol 1,4,5-trisphosphate, but only in the presence of ATP, and 32p was incorporated into it from [y-32P]ATP. There also was coincident formation of inositol 1,3,4-trisphosphate. We conclude that the more polar compound is inositol tetrakisphosphate, which is formed by phosphorylation of inositol 1,4,5-trisphosphate and may be the precursor of inositol 1,3,4-trisphosphate.Activation of several receptor-mediated cellular systems has been shown to be calcium dependent and mediated by breakdown products of polyphosphate derivatives of phosphatidylinositol (1-6). In these systems, stimulation of the relevant receptor leads to phosphodiesterase (phospholipase C)-catalyzed cleavage of phosphatidylinositol bisphosphate to inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] and 1,2-diacylglycerol (4, 5). In turn, Ins(1,4,5)P3 and diacylglycerol act as second messengers to release intracellular stores of calcium and activate protein kinase C (6), respectively. Antibodies directed against the T-lymphocyte receptor for nominal antigen or its closely associated structure, T3 (Leu-4) have been shown to mimic antigen (7) and lead to changes characteristic of activation. These changes include production of lymphokines (8) and interleukin-2 receptors (9) and proliferation (10, 11). In a series of papers (12-14), Stobo and colleagues have shown that perturbation of the T-cell receptor-T3 complex on cells of the human T-cell leukemia line Jurkat leads to production of inositol phosphates (14) and rapid increases in intracellular calcium concentration (12,13). Furthermore, in studies using permeabilized Jurkat cells, they have demonstrated that addition of inositol trisphosphate causes release of calcium from iniracellular stores (14).We report here that perturbation of the T-cell antigen receptor-T3 complex with a monoclonal antibody leads to the production of not only Ins(1,4,5)P3, but also a second inositol polyphosphate. We present evidence that this compound isThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact. inositol tetrakisphosphate (InsP4). Furthermore, we present evidence that it is generated by the ATP-dependent phosphorylation of Ins(1,4,5)P3. MATERIALS AND METHODSCells and Antibo...

Inositol-1,4,5-trisphosphate 3-kinase-A (ITPKA) is frequently over-expressed and functions as an oncogene in several tumor types

“…InsP 3 Kinase-activity was first measured in xenopus oocyes, rat liver, pancreas and brain [6]. In addition, Steward et al (1986) [7] detected InsP 3 Kinase activity in Jurkat T-cells. In 1991 Takazawa et al [8] were able to clone the first InsP 3 Kinase, which therefore was named InsP 3 Kinase-A (gene name: ITPKA).…”

Activation of phospholipase D: a signaling system set in motion by perturbation of the T lymphocyte antigen receptor/CD3 complex.

“…(Takahashi et al, 1989)]; anti-CD2 MAb, OKT1 1 (American Type Culture Collection, Bethesda, MD); anti-CD5 MAb, anti-Leu-1 (Becton Dickinson, Mountain View, CA); and anti-CD45 MAb, anti-Leu-1 8 (Becton Dickinson). At the end of stimulation, acidic phospholipids were extracted as described previously (Stewart et al, 1986) using the BlighDyer method (Bligh and Dyer, 1959; Measurement of DAG Cellular levels of DAG were measured by the DAG kinase assay as previously described (Bocckino et al, 1989) using the assay described by Preiss et al (1986) as modified by Wright et al (1988). Lipids were extracted from unstimulated and stimulated Jurkat cells using the Bligh-Dyer method (Bligh and Dyer, 1959) substituting H20 for the HCI used above.…”

Regulation of inositol 1,4,5-trisphosphate kinase activity after stimulation of human T cell antigen receptor.

“…Specific phosphatases can sequentially remove phosphate groups from the inositol ring, eventually converting Ins-1,4,5-P3 to free inositol (3). Alternatively, a cytoplasmic kinase that appears to have a wide tissue distribution can phosphorylate Ins-1,4,5-P3, yielding inositol 1 ,3,4,5-tetrakisphosphate (Ins-1,3,4,5-P4) (4)(5)(6). Although increases in Ins-1,3,4,5-P4 and its immediate breakdown product, inositol 1,3,4-trisphosphate (Ins-1,3,4-P3), have been observed after receptor stimulation, it is not known what role, if any, the Ins-1,3,4,5-P4 pathway plays in receptor-mediated regulation of cellular activities (4)(5)(6)(7).…”

“…Alternatively, a cytoplasmic kinase that appears to have a wide tissue distribution can phosphorylate Ins-1,4,5-P3, yielding inositol 1 ,3,4,5-tetrakisphosphate (Ins-1,3,4,5-P4) (4)(5)(6). Although increases in Ins-1,3,4,5-P4 and its immediate breakdown product, inositol 1,3,4-trisphosphate (Ins-1,3,4-P3), have been observed after receptor stimulation, it is not known what role, if any, the Ins-1,3,4,5-P4 pathway plays in receptor-mediated regulation of cellular activities (4)(5)(6)(7). At a minimum, however, the existence of two pathways for the metabolism of Ins-1,4,5-P3 suggests a mechanism for differential control of the levels of this Ca2"-mobilizing messenger.…”

“…Under appropriate conditions, however, monoclonal antibodies (MAbs) with specificity for T3/ Ti can mimic the effects of antigen and activate T lymphocytes (13). The addition of T3/Ti MAbs to Jurkat cells induces substantial increases in InsP3 and concentration ofcytoplasmic free calcium ([Ca2+]i) and stimulates the translocation of protein kinase C activity from the cytosol to a membrane fraction (8)(9)(10)(11) (6). As it has not been demonstrated that phosphorylation ofIns-1,4,5-P3 occurs on the 3 position under these conditions, we will refer to this product as simply InsP4.…”