Cell-permeant caged InsP3 ester shows that Ca2+ spike frequency can optimize gene expression

Inositol 1,4,5-trisphosphate (InsP3) releases calcium from intracellular stores and triggers complex waves and oscillations in levels of cytosolic free calcium,. To determine which longer-term responses are controlled by oscillations in InsP3 and cytosolic free calcium, it would be useful to deliver exogenous InsP3, under spatial and temporal control, into populations of unpermeabilized cells. Here we report the 15-step synthesis of a membrane-permeant, caged InsP3 derivative from myo-inositol. This derivative diffused into intact cells and was hydrolysed to produce a caged, metabolically stable InsP3 derivative. This latter derivative accumulated in the cytosol at concentrations of hundreds of micromolar, without activating the InsP3 receptor. Ultraviolet illumination uncaged an InsP3 analogue nearly as potent as real InsP3, and generated spikes of cytosolic free calcium, and stimulated gene expression via the nuclear factor of activated T cells,. The same total amount of InsP3 analogue elicited much more gene expression when released by repetitive flashes at 1-minute intervals than when released at 0.5- or ⩾2-minute intervals, as a single pulse, or as a slow sustained plateau. Thus, oscillations in cytosolic free calcium levels at roughly physiological rates maximize gene expression for a given amount of InsP3.

[1]  G. Zlokarnik,et al.  Quantitation of transcription and clonal selection of single living cells with beta-lactamase as reporter. , 1998, Science.

[2]  C. A. Parker,et al.  A new sensitive chemical actinometer - II. Potassium ferrioxalate as a standard chemical actinometer , 1956, Proceedings of the Royal Society of London. Series A. Mathematical and Physical Sciences.

[3]  M. Yoshida,et al.  Roles for hydroxyl groups of D-myo-inositol 1,4,5-trisphosphate in the recognition by its receptor and metabolic enzymes. , 1993, The Journal of biological chemistry.

[4]  G. Crabtree,et al.  Characterization of T cell mutants with defects in capacitative calcium entry: genetic evidence for the physiological roles of CRAC channels , 1995, The Journal of cell biology.

[5]  R. Lewis,et al.  Mitogen-regulated Ca2+ current of T lymphocytes is activated by depletion of intracellular Ca2+ stores. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[6]  G. Crabtree,et al.  The actions of cyclosporin A and FK506 suggest a novel step in the activation of T lymphocytes. , 1990, The EMBO journal.

[7]  G. Crabtree,et al.  Characterization of antigen receptor response elements within the interleukin-2 enhancer , 1988, Molecular and cellular biology.

[8]  A. Trautmann,et al.  Imaging early steps of human T cell activation by antigen-presenting cells. , 1992, Journal of immunology.

[9]  J. Keizer,et al.  Effect of Ca2+ influx on intracellular free Ca2+ responses in antigen-stimulated RBL-2H3 cells. , 1996, The American journal of physiology.

[10]  G. Crabtree,et al.  Signal transmission between the plasma membrane and nucleus of T lymphocytes. , 1994, Annual review of biochemistry.

[11]  T. Meyer,et al.  Reversible Desensitization of Inositol Trisphosphate-induced Calcium Release Provides a Mechanism for Repetitive Calcium Spikes* , 1996, The Journal of Biological Chemistry.

[12]  R Y Tsien,et al.  Photochemically generated cytosolic calcium pulses and their detection by fluo-3. , 1989, The Journal of biological chemistry.

[13]  M. McCloskey,et al.  FcεRI-mediated Induction of Nuclear Factor of Activated T-cells (*) , 1995, The Journal of Biological Chemistry.

[14]  R. Dolmetsch,et al.  Signaling between intracellular Ca2+ stores and depletion-activated Ca2+ channels generates [Ca2+]i oscillations in T lymphocytes , 1994, The Journal of general physiology.

[15]  J. Huff,et al.  The total synthesis of myo-inositol polyphosphates , 1989 .

[16]  R. Douglas Fields,et al.  Action Potential-Dependent Regulation of Gene Expression: Temporal Specificity in Ca2+, cAMP-Responsive Element Binding Proteins, and Mitogen-Activated Protein Kinase Signaling , 1997, The Journal of Neuroscience.

[17]  M. Berridge.  Inositol trisphosphate and calcium signalling , 1993, Nature.

[18]  M. Berridge,et al.  Cytosolic calcium oscillators , 1988, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[19]  C. Fewtrell.  Ca2+ oscillations in non-excitable cells. , 1993, Annual review of physiology.

[20]  G. Crabtree,et al.  Identification of calcineurin as a key signalling enzyme in T-lymphocyte activation , 1992, Nature.

[21]  J. Kuchtey,et al.  Subcloning the RBL‐2H3 mucosal mast cell line reduces Ca2+ response heterogeneity at the single‐cell level , 1996, Journal of cellular physiology.

[22]  G. Nolan,et al.  Single cell assay of a transcription factor reveals a threshold in transcription activated by signals emanating from the T-cell antigen receptor. , 1990, Genes & development.

[23]  Keli Xu,et al.  Calcium oscillations increase the efficiency and specificity of gene expression , 1998, Nature.

[24]  J. Putney,et al.  Spatial and temporal aspects of cellular calcium signaling , 1996, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[25]  D. Metcalfe,et al.  Detection of intracytoplasmic cytokine using flow cytometry and directly conjugated anti-cytokine antibodies. , 1995, Journal of immunological methods.

[26]  M. McCloskey,et al.  Fc epsilon RI-mediated induction of nuclear factor of activated T-cells. , 1995, The Journal of biological chemistry.

[27]  K. Leclair,et al.  Induction of IL-8 expression in T cells uses the CD28 costimulatory pathway. , 1994, Journal of immunology.

[28]  Christopher C. Goodnow,et al.  Differential activation of transcription factors induced by Ca2+ response amplitude and duration , 1997, Nature.

[29]  G. Crabtree,et al.  A 275 basepair fragment at the 5' end of the interleukin 2 gene enhances expression from a heterologous promoter in response to signals from the T cell antigen receptor , 1987, The Journal of experimental medicine.

[30]  Stuart L. Schreiber,et al.  Calcineurin is a common target of cyclophilin-cyclosporin A and FKBP-FK506 complexes , 1991, Cell.

[31]  G. Crabtree,et al.  Rapid shuttling of NF-AT in discrimination of Ca2+ signals and immunosuppression , 1996, Nature.

[32]  N. Spitzer,et al.  Distinct aspects of neuronal differentiation encoded by frequency of spontaneous Ca2+ transients , 1995, Nature.

[33]  M. Cahalan,et al.  Potassium and calcium channels in lymphocytes. , 1995, Annual review of immunology.

[34]  R Y Tsien,et al.  Controlling cell chemistry with caged compounds. , 1993, Annual review of physiology.

[35]  E. Price,et al.  Role of kinases and the phosphatase calcineurin in the nuclear shuttling of transcription factor NF-AT4 , 1996, Nature.

[36]  岡本 秀一.  The interleukin-8 AP-1 and κB-like sites are genetic end targets of FK506-sensitive pathway accompanied by calcium mobilization , 1994 .

[37]  M. Cahalan,et al.  Mitogen-induced oscillations of cytosolic Ca2+ and transmembrane Ca2+ current in human leukemic T cells. , 1989, Cell regulation.

[38]  R Y Tsien,et al.  Calcium channels, stores, and oscillations. , 1990, Annual review of cell biology.

[39]  M. Hirata,et al.  Stereospecific recognition of inositol 1,4,5-trisphosphate analogs by the phosphatase, kinase, and binding proteins. , 1990, The Journal of biological chemistry.

[40]  A. Kozikowski,et al.  Synthesis of 1D-3-deoxy-, 1D-2,3-dideoxy-, and 1D-2,3,6-trideoxy-myo-inositol 1,4,5-trisphosphate from quebrachitol, their binding affinities, and calcium release activity , 1993 .

[41]  G. Hajnóczky,et al.  The inositol trisphosphate calcium channel is inactivated by inositol trisphosphate , 1994, Nature.

[42]  J. Feeney,et al.  Photolabile precursors of inositol phosphates. Preparation and properties of 1-(2-nitrophenyl)ethyl esters of myo-inositol 1,4,5-trisphosphate. , 1989, Biochemistry.

[43]  G. Zlokarnik,et al.  Cell-permeant caged InsP 3 estershowsthat Ca 2 + spike frequency canoptimizegeneexpression , 2022 .

[44]  R Jacob,et al.  Calcium oscillations in electrically non-excitable cells. , 1990, Biochimica et biophysica acta.

[45]  P. Negulescu,et al.  Intracellular calcium dependence of gene expression in single T lymphocytes. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[46]  A. Rao.  NF-ATp: a transcription factor required for the co-ordinate induction of several cytokine genes. , 1994, Immunology today.