RAPAMYCIN SELECTIVELY BLOCKS INTERLEUKIN-2-INDUCED PROLIFERATING CELLNUCLEAR ANTIGEN GENE-EXPRESSION IN T-LYMPHOCYTE

The macrolide rapamycin arrests T lymphocytes stimulated by interleuki n-2 (IL-2) at G(1)/S. We have recently found that IL-2 induced an incr ease in the binding of discrete transcription factors of the ATF/cAMP- responsive element binding factor (CREB) family at G(1)/S, and that th is effect was inhibited by rapamycin (Feuerstein, N., Huang, D., Hinri chs, S. H., Orten, D. J., Aiyar, N., and Prystowsky, M. B. (1995) J. I mmunol, 154, 68-79), We now show, by using high resolution two-dimensi onal gel electrophoresis, that rapamycin inhibited selectively the syn thesis of three discrete IL-2-induced soluble proteins (35 kDa/pI simi lar to 5, 68 kDa/pI similar to 4, 110 kDa/pI similar to 4.3), Analysis of nuclear proteins demonstrated that rapamycin selectively blocked t he expression of proliferating cell nuclear antigen (PCNA), an obligat e cofactor of DNA polymerase-delta, an important component for DNA rep lication, Rapamycin inhibited the IL-2-induced PCNA mRNA, and the muri ne PCNA promoter activity in IL-2-stimulated cells, Inducible CRE-bind ing proteins were shown previously to be required for PCNA promoter ac tivity in IL-2-stimulated T lymphocytes, Using DNA binding gel mobilit y shift assay we demonstrated that rapamycin potently inhibited the bi nding 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 me chanism by which rapamycin inhibits PCNA gene expression may involve t he inhibition of the interaction of CREB/ATF transcription factors wit h CRE elements in the proximal PCNA promoter.