Jennifer Shaw

Induction, suppression and superinduction of lymphokine mRNA in T lymphocytes

The expression of several lymphokine gene is characterized by a common pattern of induction, suppression and superinduction. This pattern was studied at the level of cellular mRNA in the mouse T-lymphoma cell line EL4, the human T-leukemia line Jurkat and in normal human peripheral blood lymphocytes. Lymphokine mRNA was induced by stimulating the cells with the phorbol diester PMA (TPA), with or without T-lymphocyte mitogens. The induction of Interleukin-2, Interferon gamma and the Colony Stimulating Factor for granulocytes and macrophages was suppressed by Cyclosporin A at moderate concns. Furthermore, these mRNAs accumulated to extraordinarily high levels (superinduction) if the protein synthesis inhibitor cycloheximide was added during transcription. Superinduction was not due to an increased rate of transcription. CsA interrupted ongoing transcription of IL2 by a mechanism not dependent on the induction of a new protein. The co-ordinate regulation of these genes strongly suggests that common intracellular signals mediate their expression.

Cellular Origins and Targets of Costimulator (IL2)

ABBREVIATIONS: A cell cells adherent to nyton wool AEF allogeneic effect factor Con A concanavalin A CTL cytotoxic T lymphocyte IEF isoelectric focusing ILI, IL2Interleukins 1 and 2 LAF lymphocyte activating factor, same as ILI MLC mixed leukocyte culture MW molecular weight PFC plaque forming cells in an antibody response SRBC sheep red blood cells TCGF T cell growth factor, same as IL2 TMF thymocyte mitogenic factor, same as IL2 TRF T cell replacing factor TSF thymocyte stimulating factor, same as IL2

Teleocidin and phorbol ester tumor promoters exert similar mitogenic effects on human lymphocytes

The tumor promoter 12-0-tetradecanoyl-phorbol-13-acetate (TPA) affects a wide variety of cellular functions via its binding to protein kinase C (PKC). The TPA molecule contains a diacylglycerol (DAG)-like structure, which may explain its ability to mimic DAG in PKC activation. Teleocidin (TCD) is a different tumor promoter which can compete with TPA in binding to its cell surface receptors even though structurally unrelated to TPA or DAG. Since TCD may use an additional receptor system and/or be distinguished from TPA in its effect on cells, we compared the effects of TPA and TCD on human peripheral blood lymphocytes (PBL). Both tumor promoters preferentially enhanced cell proliferation of sheep erythrocyte-rosetted lymphocytes, which were enriched for T cells. Additionally, TPA and TCD both induced a high density of cell surface receptors for interleukin 2 (IL2) and transferrin, but not synthesis or production of IL2. However, either of the tumor promoters synergized with T cell mitogens to induce high level IL2 production by PBL. In dose response and kinetic studies, matching concentrations of TPA and TCD induced similar effects in PBL. The results thus demonstrate that TPA and TCD are alike in mitogenic capacity, and suggest that structural similarity between the tumor promoter and DAG, the physiological activator of PKC, is not an essential property for promoting tumors or affecting a wide variety of cellular functions.

A Molecular‐Genetic Analysis of Cytotoxic T Lymphocyte Function

Two genes that are specifically expressed in T cells with cytolytic activity were isolated from a CTL cDNA library by differential screening. Both appear to encode serine proteases, thus suggesting a cascade mechanism, similar to complement, in activated CTL. Both CTL-specific proteases have a number of unusual structural features that suggest that they will have novel substrate specificities. One of the proteins (CCPI) has been oriented to the granules found in the cytoplasm of CTL. Taken together, these data strongly suggest that these molecules play an important role in target-cell lysis by CTL. Furthermore, we believe that the detailed molecular knowledge being accumulated through these studies may lead to the development of innovative forms of immunotherapy.

Life after IL2

Upon interaction with interleukin 2 and antigen, a precursor cyto - toxic T lymphocyte (pCTL) undergoes the final steps of differentiation to become an activated killer cell capable of binding to and lysing antigen bearing target cells. In response to these stimuli, a variety of genes are upregulated including those which encode the IL2-receptor (Kronke et al., 1985), the receptors for transferrin and insulin (Neckers and Cossman, 1983), the c-myc protein (Kelly et al., 1983), and MHC molecules (Cotner et al., 1983). These gene products play important roles in cell growth, cycling and communication, however they are not directly involved in the cytolytic machinery of activated CTL.

Regulation of IL2 and Related Genes at the mRNA Level

Synthesis and secretion of lymphokines is highly regulated in normal T lymphocytes and in several human and murine T lymphocyte cell lines. The responses of the cell lines serve as useful models for the induction of primary T helper lymphocytes, since the two types of cells share a number of features. For example, the phorbol diester PMA (TPA) is a co-stimulator for the induction of IL2 and other lymphokines in both primary cells (1) and cell lines (2,3). Furthermore, in both types of cells, the induction of IL2 protein synthesis is due to a rapid rise in the level of its mRNA (4,5,6,7), and the same is true of certain other lymphokines such as GM-CSF (6) and interferon gamma (8). Induction of lymphokines in the mouse T lymphocyte cell line EL4 by PMA (2) is not accompanied by general alterations in gene expression (9), rather, it reflects a specific enhancement of transcription of only a small set of genes, including several lymphokines (6).