Gene array and protein expression profiles suggest post-transcriptional regulation during CD8+ T cell differentiation.
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.
Cham
Candace M
CM
Committee on Cancer Biology, University of Chicago, Chicago, Illinois 60637, USA.
Xu
Hui
H
O'Keefe
James P
JP
Rivas
Fabiola V
FV
Zagouras
Panayiotis
P
Gajewski
Thomas F
TF
D000818:Animals; D016827:CD8 Antigens; D018414:CD8-Positive T-Lymphocytes / Q000502:physiology*; D002454:Cell Differentiation / Q000502:physiology*; D016207:Cytokines / Q000096:biosynthesis* / Q000235:genetics; D015536:Down-Regulation; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D051379:Mice; D020411:Oligonucleotide Array Sequence Analysis; D012313:RNA / Q000032:analysis / Q000235:genetics; D012323:RNA Processing, Post-Transcriptional; D015854:Up-Regulation
D016827:CD8 Antigens; D016207:Cytokines; D012313:RNA