A soluble form of CTLA‐4 generated by alternative splicing is expressed by nonstimulated human T cells

CTLA‐4, expressed by activated T cells, transduces an inhibitory signal. We show here that PCR amplification of the coding sequence of CTLA‐4 in nonstimulated human T lymphocytes results in the amplification of two transcripts of 650 and 550 bp. Sequencing shows that the larger form codes for membrane CTLA‐4 and the 550‐bp transcript is a spliced variant in which exon 2 coding for the transmembrane region is deleted. This spliced cDNA has been named CTLA‐4delTM. The splicing induces a frame shift which results in the addition of 22 extra amino acids before a translational termination. Activation of T cells with phorbol 12‐myristate 13‐acetate plus ionomycin or anti‐CD3 plus anti‐CD28 monoclonal antibodies induces a suppression of CTLA‐4delTM mRNA expression associated with a preferential expression of the membrane CTLA‐4 mRNA, showing that CTLA‐4delTM mRNA expression is restricted to nonactivated T cells. A soluble immunoreactive form of CTLA‐4 was detected in the serum of 14 / 64 healthy subjects. These results suggest that nonstimulated T cells may constitutively produce a soluble form of CTLA‐4 which may have an important role in the regulation of immune homeostasis.

[1]  Stephen M. Mount,et al.  A catalogue of splice junction sequences. , 1982, Nucleic acids research.

[2]  R. Karr,et al.  Antigen-dependent clonal expansion of a trace population of antigen-specific CD4+ T cells in vivo is dependent on CD28 costimulation and inhibited by CTLA-4. , 1995, Journal of immunology.

[3]  J. Allison,et al.  CD28 and CTLA-4 have opposing effects on the response of T cells to stimulation , 1995, The Journal of experimental medicine.

[4]  P. Linsley,et al.  CD80 (B7-1) Binds Both CD28 and CTLA-4 with a Low Affinity and Very Fast Kinetics , 1997, The Journal of experimental medicine.

[5]  P. Linsley,et al.  Binding of the B cell activation antigen B7 to CD28 costimulates T cell proliferation and interleukin 2 mRNA accumulation , 1991, The Journal of experimental medicine.

[6]  P. Linsley,et al.  The role of the CD28 receptor during T cell responses to antigen. , 1993, Annual review of immunology.

[7]  J. Allison,et al.  Enhancement of Antitumor Immunity by CTLA-4 Blockade , 1996, Science.

[8]  R. Karr,et al.  Long-term inhibition of murine experimental autoimmune encephalomyelitis using CTLA-4-Fc supports a key role for CD28 costimulation. , 1995, The Journal of clinical investigation.

[9]  T. Saito.  Negative regulation of T cell activation. , 1998, Current opinion in immunology.

[10]  J. Bluestone,et al.  Loss of CTLA-4 leads to massive lymphoproliferation and fatal multiorgan tissue destruction, revealing a critical negative regulatory role of CTLA-4. , 1995, Immunity.

[11]  Z. Wang,et al.  Synergistic induction of CTLA-4 expression by costimulation with TCR plus CD28 signals mediated by increased transcription and messenger ribonucleic acid stability. , 1997, Journal of immunology.

[12]  L. Lanier,et al.  CD28 interaction with B7 costimulates primary allogeneic proliferative responses and cytotoxicity mediated by small, resting T lymphocytes , 1992, The Journal of experimental medicine.

[13]  B. Hausmann,et al.  Mice transgenic for a soluble form of murine CTLA-4 show enhanced expansion of antigen-specific CD4+ T cells and defective antibody production in vivo , 1994, The Journal of experimental medicine.

[14]  J. Allison,et al.  CD28-B7 interactions allow the induction of CD8+ cytotoxic T lymphocytes in the absence of exogenous help , 1993, The Journal of experimental medicine.

[15]  J. Bluestone,et al.  The B7 and CD28 receptor families. , 1994, Immunology today.

[16]  L. Maiuri,et al.  Blockage of T-cell costimulation inhibits T-cell action in celiac disease. , 1998, Gastroenterology.

[17]  P. Linsley,et al.  Binding Stoichiometry of the Cytotoxic T Lymphocyte-associated Molecule-4 (CTLA-4) , 1995, The Journal of Biological Chemistry.

[18]  P. Linsley,et al.  Complementarity determining region 1 (CDR1)- and CDR3-analogous regions in CTLA-4 and CD28 determine the binding to B7-1 , 1994, The Journal of experimental medicine.

[19]  M. Lefranc,et al.  Human Ig superfamily CTLA‐4 gene: chromosomal localization and identity of protein sequence between murine and human CTLA‐4 cytoplasmic domains , 1988, European journal of immunology.

[20]  J. Bluestone,et al.  Regulation of CTLA-4 expression during T cell activation. , 1996, Journal of immunology.

[21]  J. Bonifacino,et al.  Tyrosine phosphorylation controls internalization of CTLA-4 by regulating its interaction with clathrin-associated adaptor complex AP-2. , 1997, Immunity.

[22]  P. Linsley,et al.  Covalent Dimerization of CD28/CTLA-4 and Oligomerization of CD80/CD86 Regulate T Cell Costimulatory Interactions* , 1996, The Journal of Biological Chemistry.

[23]  M. Mattei,et al.  CTLA-4 and CD28 activated lymphocyte molecules are closely related in both mouse and human as to sequence, message expression, gene structure, and chromosomal location. , 1991, Journal of immunology.

[24]  J. Allison,et al.  Co-stimulation in T cell responses. , 1997, Current opinion in immunology.

[25]  K. Kohno,et al.  CD28 molecule as a receptor-like function for accessory signals in cell-mediated augmentation of IL-2 production. , 1990, Cellular immunology.

[26]  F. Denizot,et al.  A new member of the immunoglobulin superfamily—CTLA-4 , 1987, Nature.

[27]  G. Freeman,et al.  Characterization of CTLA-4 structure and expression on human T cells. , 1993, Journal of immunology.

[28]  M. Feldmann,et al.  Prevention and amelioration of collagen‐induced arthritis by blockade of the CD28 co‐stimulatory pathway: requirement for both B7‐1 and B7‐2 , 1996, European journal of immunology.

[29]  J. Gribben,et al.  B-cell surface antigen B7 provides a costimulatory signal that induces T cells to proliferate and secrete interleukin 2. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[30]  P. Linsley,et al.  CTLA-4 is a second receptor for the B cell activation antigen B7 , 1991, The Journal of experimental medicine.

[31]  K P Lee,et al.  Differential T cell costimulatory requirements in CD28-deficient mice. , 1993, Science.

[32]  P. Linsley,et al.  Treatment of murine lupus with CTLA4Ig. , 1994, Science.

[33]  H. Griesser,et al.  Lymphoproliferative Disorders with Early Lethality in Mice Deficient in Ctla-4 , 1995, Science.

[34]  P. Linsley,et al.  CTLA-4 can function as a negative regulator of T cell activation. , 1994, Immunity.

[35]  M. V. Vander Heiden,et al.  Interaction of CTLA-4 with the clathrin-associated protein AP50 results in ligand-independent endocytosis that limits cell surface expression. , 1997, Journal of immunology.

[36]  J. Allison,et al.  Specific blockade of CTLA-4/B7 interactions results in exacerbated clinical and histologic disease in an actively-induced model of experimental allergic encephalomyelitis , 1997, Journal of Neuroimmunology.

[37]  P. Linsley,et al.  Direct helper T cell‐induced B cell differentiation involves interaction between T cell antigen CD28 and B cell activation antigen B7 , 1991, European journal of immunology.

[38]  G. Freeman,et al.  CTLA-4 and CD28 mRNA are coexpressed in most T cells after activation. Expression of CTLA-4 and CD28 mRNA does not correlate with the pattern of lymphokine production. , 1992, Journal of immunology.

[39]  P. Linsley,et al.  Immunosuppression in vivo by a soluble form of the CTLA-4 T cell activation molecule. , 1992, Science.

[40]  R. Schwartz,et al.  A cell culture model for T lymphocyte clonal anergy. , 1990, Science.

[41]  I. Kariv,et al.  Analysis of the site of interaction of CD28 with its counter-receptors CD80 and CD86 and correlation with function. , 1996, Journal of immunology.