Comparison of T Cell Receptor-Induced Proximal Signaling and Downstream Functions in Immortalized and Primary T Cells

Abstract

BackgroundHuman T cells play an important role in pathogen clearance, but their aberrant activation is also linked to numerous diseases. T cells are activated by the concurrent induction of the T cell receptor (TCR) and one or more costimulatory receptors. The characterization of signaling pathways induced by TCR and/or costimulatory receptor activation is critical, since these pathways are excellent targets for novel therapies for human disease. Although studies using human T cell lines have provided substantial insight into these signaling pathways, no comprehensive, direct comparison of these cell lines to activated peripheral blood T cells (APBTs) has been performed to validate their usefulness as a model of primary T cells.Methodology/Principal FindingsWe used quantitative biochemical techniques to compare the activation of two widely used human T cell lines, Jurkat E6.1 and HuT78 T cells, to APBTs. We found that HuT78 cells were similar to APBTs in proximal TCR-mediated signaling events. In contrast, Jurkat E6.1 cells had significantly increased site-specific phosphorylation of Pyk2, PLCγ1, Vav1, and Erk1/Erk2 and substantially more Ca2+ flux compared to HuT78 cells and APBTs. In part, these effects appear to be due to an overexpression of Itk in Jurkat E6.1 cells compared to HuT78 cells and APBTs. Both cell lines differ from APBTs in the expression and function of costimulatory receptors and in the range of cytokines and chemokines released upon TCR and costimulatory receptor activation.Conclusions/SignificanceBoth Jurkat E6.1 and HuT78 T cells had distinct similarities and differences compared to APBTs. Both cell lines have advantages and disadvantages, which must be taken into account when choosing them as a model T cell line.

An analysis of the phosphoproteome of immune cell lines exposed to the immunomodulatory mycotoxin deoxynivalenol

“…The use of immortalized cell lines has several advantages such as the cells being easy to grow and maintain, provide unlimited experimental material, among others. Although it has been documented that Jurkat E6.1 cells have specific abnormalities [18], they have been widely and successfully used in signaling transduction investigation [20] to reveal real, normal events which were further validated in other models [18,21]. RPMI1788 cells have also been used in several signaling transduction studies [22], although due to their primary origin these cells do not raise the same objections as Jurkat E6.1 cells.…”

“…We now extend this work to compare phosphoprotein expression in human B (RPMI1788) and T (Jurkat E6.1) lymphocyte cell lines exposed to DON at a maximum concentration of 500 ng DON/mL for a period of up to 54 h. These cell types represent major cellular components of the adaptive immune response and have been used extensively previously [17,18]. A panel of phosphoproteins which exhibited validated altered phosphorylation state across these cell lines is described.…”

Characterization of dynamic actin associations with T-cell receptor microclusters in primary T cells

“…This might add to already-documented differences in membrane protein organization between Jurkat cells and primary T cells (Tanimura et al, 2003). Furthermore, a number of Jurkat cell proteins exhibit levels of phosphorylation unlike those of their primary T cell counterparts, including known actin regulatory molecules such as Pyk2 and Vav1 (Bartelt et al, 2009). These differences raise questions about findings from studies based on Jurkat cell studies and necessitate the ultimate characterization of actin cytoskeleton behavior in primary cells.…”

“…Preliminary observations of actin engaged in such interactions have been made in Jurkat cells (Kaizuka et al, 2007;Yu et al, 2010), but although these are a tractable T-cell-based model system, they have important differences from primary T cells (Tanimura et al, 2003;Bartelt et al, 2009;Dobson-Belaire et al, 2011). Because their TCR agonist is not known, Jurkat cells must be triggered through artificial antibody-induced crosslinking of their CD3 receptors, which inherently supplants endogenous TCR-TCR associations and has the potential to overstabilize the microcluster.…”

Copy Number Networks to Guide Combinatorial Therapy of Cancer and Proliferative Disorders

“…This gene plays a key role in T cell activation, an important cellular response in autoimmune disorders (Bartelt et al, 2009;Kannan et al, 2012;SmithGarvin et al, 2009). In the RH survival network, 56 genes were linked with statistical significance (FDR < 10 -4 ) to NFATc1 ( Figure 2C).…”