Rapamycin selectively blocks interleukin-2-induced proliferating cell nuclear antigen gene expression in T lymphocyte. Evidence for inhibition of CREB/ATF binding activities.
The macrolide rapamycin arrests T lymphocytes stimulated by interleukin-2 (IL-2) at G1/S. We have recently found that IL-2 induced an increase in the binding of discrete transcription factors of the ATF/cAMP-responsive element binding factor (CREB) family at G1/S, and that this effect was inhibited by rapamycin (Feuerstein, N., Huang, D., Hinrichs, S. H., Orten, D. J., Aiyar, N., and Prystowsky, M. B. (1995) J. Immunol. 154, 68-79). We now show, by using high resolution two-dimensional gel electrophoresis, that rapamycin inhibited selectively the synthesis of three discrete IL-2-induced soluble proteins (35 kDa/pI approximately 5, 68 kDa/pI approximately 4, 110 kDa/pI approximately 4.3). Analysis of nuclear proteins demonstrated that rapamycin selectively blocked the expression of proliferating cell nuclear antigen (PCNA), an obligate cofactor of DNA polymerase-delta, an important component for DNA replication. Rapamycin inhibited the IL-2-induced PCNA mRNA, and the murine PCNA promoter activity in IL-2-stimulated cells. Inducible CRE-binding proteins were shown previously to be required for PCNA promoter activity in IL-2-stimulated T lymphocytes. Using DNA binding gel mobility shift assay we demonstrated that rapamycin potently inhibited the binding of CREB/ATF transcription factors to CRE elements in the murine proximal PCNA promoter. These results suggest that PCNA is a preferred target in a rapamycin-sensitive transduction pathway, and that the mechanism by which rampamycin inhibits PCNA gene expression may involve the inhibition of the interaction of CREB/ATF transcription factors with CRE elements in the proximal PCNA promoter.
Feuerstein
N
N
Medical College of Pennsylvania, Philadelphia 19129, USA.
Huang
D
D
Prystowsky
M B
MB
D051696:Activating Transcription Factors; D000818:Animals; D001483:Base Sequence; D001798:Blood Proteins / Q000037:antagonists & inhibitors*; D002478:Cells, Cultured; D017362:Cyclic AMP Response Element-Binding Protein / Q000037:antagonists & inhibitors*; D005786:Gene Expression Regulation / Q000187:drug effects*; D007376:Interleukin-2 / Q000494:pharmacology*; D051379:Mice; D008969:Molecular Sequence Data; D011090:Polyenes / Q000494:pharmacology*; D018809:Proliferating Cell Nuclear Antigen / Q000235:genetics*; D011401:Promoter Regions, Genetic; D020123:Sirolimus; D013601:T-Lymphocytes / Q000187:drug effects* / Q000378:metabolism; D014157:Transcription Factors / Q000037:antagonists & inhibitors*; D014158:Transcription, Genetic
D051696:Activating Transcription Factors; D001798:Blood Proteins; D017362:Cyclic AMP Response Element-Binding Protein; D007376:Interleukin-2; D011090:Polyenes; D018809:Proliferating Cell Nuclear Antigen; D014157:Transcription Factors; D020123:Sirolimus