IFN-γ upregulates survivin and Ifi202 expression to induce survival and proliferation of tumor-specific T cells.
BACKGROUND
A common procedure in human cytotoxic T lymphocyte (CTL) adoptive transfer immunotherapy is to expand tumor-specific CTLs ex vivo using CD3 mAb prior to transfer. One of the major obstacles of CTL adoptive immunotherapy is a lack of CTL persistence in the tumor-bearing host after transfer. The aim of this study is to elucidate the molecular mechanisms underlying the effects of stimulation conditions on proliferation and survival of tumor-specific CTLs.


RESULTS
Tumor-specific CTLs were stimulated with either CD3 mAb or cognate Ag and analyzed for their proliferation and survival ex vivo and persistence in tumor-bearing mice. Although both Ag and CD3 mAb effectively induced the cytotoxic effecter molecules of the CTLs, we observed that Ag stimulation is essential for sustained CTL proliferation and survival. Further analysis revealed that Ag stimulation leads to greater proliferation rates and less apoptosis than CD3 mAb stimulation. Re-stimulation of the CD3 mAb-stimulated CTLs with Ag resulted in restored CTL proliferative potential, suggesting that CD3 mAb-induced loss of proliferative potential is reversible. Using DNA microarray technology, we identified that survivin and ifi202, two genes with known functions in T cell apoptosis and proliferation, are differentially induced between Ag- and CD3 mAb-stimulated CTLs. Analysis of the IFN-γ signaling pathway activation revealed that Ag stimulation resulted in rapid phosphorylation of STAT1 (pSTAT1), whereas CD3 mAb stimulation failed to activate STAT1. Chromatin immunoprecipitation revealed that pSTAT1 is associated with the promoters of both survivin and ifi202 in T cells and electrophoresis mobility shift assay indicated that pSTAT1 directly binds to the gamma activation sequence element in the survivin and ifi202 promoters. Finally, silencing ifi202 expression significantly decreased T cell proliferation.


CONCLUSIONS
Our findings delineate a new role of the IFN-γ signaling pathway in regulating T cell proliferation and apoptosis through upregulating survivin and ifi202 expression.
Department of Biochemistry and Molecular Biology, Medical College of Georgia, Georgia Health Sciences University, Augusta, Georgia, USA.
Mary
M
Zimmerman
Dafeng
D
Yang
Xiaolin
X
Hu
Feiyan
F
Liu
Nagendra
N
Singh
Darren
D
Browning
Vadivel
V
Ganapathy
Phillip
P
Chandler
Divaker
D
Choubey
Scott I
SI
Abrams
Kebin
K
Liu
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D017252:CD3 Complex / Q000276:immunology; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D005260:Female; D020869:Gene Expression Profiling; D016219:Immunotherapy, Adoptive; D051034:Inhibitor of Apoptosis Proteins / Q000235:genetics / Q000378:metabolism*; D007371:Interferon-gamma / Q000378:metabolism* / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D009374:Neoplasms, Experimental / Q000378:metabolism / Q000473:pathology / Q000628:therapy; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D034622:RNA Interference; D017471:Receptors, Interferon / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D050794:STAT1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D000077022:Survivin; D013602:T-Lymphocytes, Cytotoxic / Q000166:cytology / Q000378:metabolism* / Q000637:transplantation; D015854:Up-Regulation / Q000187:drug effects; D000096743:Interferon gamma Receptor
D000911:Antibodies, Monoclonal; D017252:CD3 Complex; D051034:Inhibitor of Apoptosis Proteins; D007371:Interferon-gamma; D047908:Intracellular Signaling Peptides and Proteins; D017471:Receptors, Interferon; D012097:Repressor Proteins; D050794:STAT1 Transcription Factor; D000077022:Survivin; D000096743:Interferon gamma Receptor; C117749:Birc5 protein, mouse; C092537:Ifi202b protein, mouse; C494080:Stat1 protein, mouse