As targets for the immunosuppressive drugs cyclosporin A and FK506, transcription factors of the NFAT (nuclear factor of activated T cells) family have been the focus of much attention. NFAT proteins, which are expressed in most immune-system cells, play a pivotal role in the transcription of cytokine genes and other genes critical for the immune response. The activity of NFAT proteins is tightly regulated by the calcium/calmodulin-dependent phosphatase calcineurin, a primary target for inhibition by cyclosporin A and FK506. Calcineurin controls the translocation of NFAT proteins from the cytoplasm to the nucleus of activated cells by interacting with an N-terminal regulatory domain conserved in the NFAT family. The DNA-binding domains of NFAT proteins resemble those of Rel-family proteins, and Rel and NFAT proteins show some overlap in their ability to bind to certain regulatory elements in cytokine genes. NFAT is also notable for its ability to bind cooperatively with transcription factors of the AP-1 (Fos/Jun) family to composite NFAT:AP-1 sites, found in the regulatory regions of many genes that are inducibly transcribed by immune-system cells. This review discusses recent data on the diversity of the NFAT family of transcription factors, the regulation of NFAT proteins within cells, and the cooperation of NFAT proteins with other transcription factors to regulate the expression of inducible genes.

TRANSCRIPTION FACTORS OF THE NFAT FAMILY:Regulation and Function

The importance of the B7/CD28/CTLA-4 molecules has been established in studies of antigen-presenting cell-derived B7 and its interaction with the T cell costimulatory molecule CD28. CTLA-4, a T cell surface glycoprotein that is related to CD28, can also interact with B7-1 and B7-2. However, less is known about the function of CTLA-4, which is expressed at highest levels after activation. We have generated an antibody to CTLA-4 to investigate the consequences of engagement of this molecule in a carefully defined system using highly purified T cells. We show here that the presence of low levels of B7-2 on freshly explanted T cells can partially inhibit T cell proliferation, and this inhibition is mediated by interactions with CTLA-4. Cross-linking of CTLA-4 together with the TCR and CD28 strongly inhibits proliferation and IL-2 secretion by T cells. Finally, results show that CD28 and CTLA-4 deliver opposing signals that appear to be integrated by the T cell in determining the response to activation. These data strongly suggest that the outcome of T cell antigen receptor stimulation is regulated by CD28 costimulatory signals, as well as inhibitory signals derived from CTLA-4.

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CD28 and CTLA-4 have opposing effects on the response of T cells to stimulation.

T lymphocytes respond to foreign antigens both by producing protein effector molecules known as lymphokines and by multiplying. Complete activation requires two signaling events, one through the antigen-specific receptor and one through the receptor for a costimulatory molecule. In the absence of the latter signal, the T cell makes only a partial response and, more importantly, enters an unresponsive state known as clonal anergy in which the T cell is incapable of producing its own growth hormone, interleukin-2, on restimulation. Our current understanding at the molecular level of this modulatory process and its relevance to T cell tolerance are reviewed.

http://science.sciencemag.org/content/sci/248/4961/1349.full.pdf

A cell culture model for T lymphocyte clonal anergy

AU-rich elements: characterization and importance in mRNA degradation.

Functional interactions between T and B lymphocytes are necessary for optimal activation of an immune response. Recently, the T lymphocyte receptor CD28 was shown to bind the B7 counter-receptor on activated B lymphocytes, and subsequently to costimulate interleukin 2 production and T cell proliferation. CTLA-4 is a predicted membrane receptor from cytotoxic T cells that is homologous to CD28 and whose gene maps to the same chromosomal band as the gene for CD28. It is not known, however, if CD28 and CTLA-4 also share functional properties. To investigate functional properties of CTLA-4, we have produced a soluble genetic fusion between the extracellular domain of CTLA-4 and an immunoglobulin C gamma chain. Here, we show that the fusion protein encoded by this construct, CTLA4Ig, bound specifically to B7-transfected Chinese hamster ovary cells and to lymphoblastoid cells. CTLA4Ig also immunoprecipitated B7 from cell surface 125I-labeled extracts of these cells. The avidity of 125I-labeled B7Ig fusion protein for immobilized CTLA4Ig was estimated (Kd approximately 12 nM). Finally, we show that CTLA4Ig was a potent inhibitor of in vitro immune responses dependent upon cellular interactions between T and B lymphocytes. These findings provide direct evidence that, like its structural homologue CD28, CTLA-4 is able to bind the B7 counter-receptor on activated B cells. Lymphocyte interactions involving the B7 counter-receptor are functionally important for alloantigen responses in vitro.

/pdf/ctla-4-is-a-second-receptor-for-the-b-cell-activation-qrg6etpfrm.pdf

CTLA-4 is a second receptor for the B cell activation antigen B7.

Quiescent T cells can be induced to express many genes by mitogen or antigen stimulation. The messenger RNAs of some of these genes undergo relatively rapid degradation compared to messenger RNAs from constitutively expressed genes. A T cell activation pathway that specifically regulates the stability of messenger RNAs for the lymphokines interleukin-2, interferon-gamma, tumor necrosis factor-alpha, and granulocyte-macrophage colony-stimulating factor is induced by stimulation of the CD28 surface molecule. This pathway does not directly affect the steady-state messenger RNA level, transcription, or messenger RNA half-life of other T cell activation genes, including c-myc, c-fos, IL-2 receptor, and the 4F2HC surface antigen. These data show that stimuli received at the cell surface can alter gene expression by inducing specific changes in messenger RNA degradation.

https://science.sciencemag.org/content/sci/244/4902/339.full.pdf

Regulation of lymphokine messenger RNA stability by a surface-mediated T cell activation pathway

The regulation of IL-3 gene induction in human peripheral blood T cells was studied. IL-3 gene expression was inducible by crosslinking of the T cell receptor/CD3 complex using anti-CD3 MAb G19-4. Anti-CD3-induced IL-3 gene expression was found to be limited to the CD28+ T cell subset and could be augmented by costimulating T lymphocytes with antibodies directed against CD28. IL-3 expression could also be induced by costimulation of T cells with both phorbol ester and ionomycin, which are thought to mimic the intracellular effects of T cell receptor-antigen interaction. However, unlike other lymphokines such as IL-2 or granulocyte-macrophage colony-stimulating factor, IL-3 gene expression is not induced by stimulation of cells with phorbol myristate acetate and anti-CD28. We conclude that IL-3 gene regulation is under stringent control since IL-3 gene expression occurs only in the CD28+ subset of T cells, and since IL-3 induction obligately requires increased intracellular calcium.

/pdf/regulation-of-interleukin-3-gene-induction-in-normal-human-t-1gy09q59zc.pdf

Regulation of interleukin 3 gene induction in normal human T cells.

Unlike mammals, chickens generate an immunoglobulin (Ig) repertoire by a developmentally regulated process of intrachromosomal gene conversion, which results in nucleotide substitutions throughout the variable regions of the Ig heavy- and light-chain genes. In contrast to chicken Ig genes, we show in this report that diversity of the rearranged chicken T-cell receptor (TCR) beta-chain gene is generated by junctional heterogeneity, as observed in rearranged mammalian TCR genes. This junctional diversity increases during chicken development as a result of an increasing base-pair addition at the V beta-D beta and D beta-J beta joints (where V, D, and J are the variable, diversity, and joining gene segments). Despite the junctional hypervariability, however, almost all functional V beta-D beta-J beta junctions appear to encode a glycine-containing beta-turn. Such a turn may serve to position the amino acid side chains of a hypervariable TCR beta-chain loop with respect to the antigen-binding groove of the major histocompatibility complex molecule. Consistent with this hypothesis, the germ-line D beta nucleotide sequences of chickens, mice, rabbits, and humans have been highly conserved and encode a glycine in all three reading frames.

https://www.pnas.org/content/pnas/88/17/7699.full.pdf

Gregory Stella

Papers

Regulation of lymphokine messenger RNA stability by a surface-mediated T cell activation pathway

Regulation of interleukin 3 gene induction in normal human T cells.

Chicken T-cell receptor beta-chain diversity: an evolutionarily conserved D beta-encoded glycine turn within the hypervariable CDR3 domain.