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We recently reported that differentiation of CD8(+) T cells from the naïve to the effector state involves the upregulation of glucose-dependent metabolism. Glucose deprivation or inhibition of glycolysis by 2-deoxy-D-glucose (2-DG) selectively inhibited production of IFN-gamma but not of IL-2. To determine a more global role of glucose metabolism on effector T-cell function, we performed gene array analysis on CD8(+) effector T cells stimulated in the presence or absence of 2-DG. We observed that expression of only 10% of genes induced by TCR/CD28 signaling was inhibited by 2-DG. Among these were genes for key cytokines, cell cycle molecules, and cytotoxic granule proteins. Consistent with these results, production of IFN-gamma and GM-CSF, cell cycle progression, upregulation of cyclin D2 protein, cytolytic activity, and upregulation of granzyme B protein and also conjugate formation were exquisitely glucose-dependent. In contrast to glucose, oxygen was little utilized by CD8(+) effector T cells, and relative oxygen deprivation did not inhibit these CTL functional properties. Our results indicate a particularly critical role for glucose in regulating specific effector functions of CD8(+) T cells and have implications for the maintenance of the effector phase of cellular immune responses in target tissue microenvironments such as a solid tumor.

Peripheral CD8(+) T cells circulate in a quiescent naive state until they are primed by specific antigen and differentiate into effector cells. In the effector state, CD8(+) T cells acquire cytolytic activity and produce increased levels of cytokines such as interferon-gamma. They also exhibit increased T cell receptor sensitivity, decreased CD28 dependence, and become inhibitable by CTLA-4 and other negative regulatory pathways. We hypothesized that one mechanism by which these two states are regulated is via differential expression of specific genes. To this end, basal gene expression profiles of naive and effector 2C TCR transgenic x RAG2(-/-) CD8(+) T cells were analyzed using Affymetrix arrays representing 11,000 genes. Of the 177 differentially expressed known genes, 68 were expressed at higher levels in effector cells, but 109 were more abundant in naive cells, supporting the notion that the naive state is not passive. Expression of genes related to metabolism, actin cytoskeletal dynamics, and effector function increased with priming, whereas expression of putative anti-proliferative genes decreased. Semiquantitative reverse transcription-PCR was utilized as a secondary validation for selected transcripts, and Western blot analysis was used to examine protein expression for molecules of interest. Surprisingly, for 24 genes examined, 12 showed discordant protein versus mRNA expression. In summary, our study indicates that: 1) not only does the expression of some genes in naive CD8(+) T cells become up-regulated upon priming, but the expression of other genes is down-regulated as well and 2) the complexities of T cell differentiation include regulation at the post-transcriptional level.