The 5' flanking region of the human interleukin (IL)-2 gene was investigated for enhancer activity in response to CD69-generated signals, using a chloramphenicol acetyltransferase (CAT)-driven transient expression system in Jurkat cells. The region extending from -317 to +47 relative to the initiation site of IL-2 gene transcription was shown to contain sequences able to respond to CD69 cross-linking, by enhancing by about 100% a phorbol 12-myristate 13-acetate (PMA)-plus-ionomycin stimulation of CAT activity. A similar increase in CAT activity produced by PMA-plus-anti-CD3 mAb was induced by CD69 crosslinking, while a 200% increase over that obtained by PMA-plus-anti-CD28 mAb stimulation was seen. Analysis of enhancer deletion mutants revealed that proximal AP-1, OCT-1/octamer-associated protein and nuclear factor of activated T cells (NFAT) binding regions were all necessary to allow CD69-mediated enhancement of CAT activity. By gel mobility shift analysis, cyclosporin A-sensitive NFAT-binding induction and enhancement of AP-1 binding activity could be detected in nuclear extracts of both Jurkat and peripheral blood T cells after simultaneous CD69 and protein kinase C stimulation. Finally, CD69-mediated signals could increase NFAT and AP-1 binding activity following PMA and ionomycin stimulation in peripheral blood T cells. Collectively, these data suggest that CD69-generated signals participate in the control of the IL-2 gene expression at the transcriptional level, likely acting through NFAT and AP-1 transcription factor complexes.
Transcriptional regulation of interleukin-2 gene expression by CD69-generated signals / D'Ambrosio, D; Trotta, R; Vacca, A; Frati, Luigi; Santoni, Angela; Gulino, Alberto; Testi, R.. - In: EUROPEAN JOURNAL OF IMMUNOLOGY. - ISSN 0014-2980. - STAMPA. - 23:(1993), pp. 2993-2997. [10.1002/eji.1830231140]
Transcriptional regulation of interleukin-2 gene expression by CD69-generated signals.
FRATI, Luigi;SANTONI, Angela;GULINO, Alberto;
1993
Abstract
The 5' flanking region of the human interleukin (IL)-2 gene was investigated for enhancer activity in response to CD69-generated signals, using a chloramphenicol acetyltransferase (CAT)-driven transient expression system in Jurkat cells. The region extending from -317 to +47 relative to the initiation site of IL-2 gene transcription was shown to contain sequences able to respond to CD69 cross-linking, by enhancing by about 100% a phorbol 12-myristate 13-acetate (PMA)-plus-ionomycin stimulation of CAT activity. A similar increase in CAT activity produced by PMA-plus-anti-CD3 mAb was induced by CD69 crosslinking, while a 200% increase over that obtained by PMA-plus-anti-CD28 mAb stimulation was seen. Analysis of enhancer deletion mutants revealed that proximal AP-1, OCT-1/octamer-associated protein and nuclear factor of activated T cells (NFAT) binding regions were all necessary to allow CD69-mediated enhancement of CAT activity. By gel mobility shift analysis, cyclosporin A-sensitive NFAT-binding induction and enhancement of AP-1 binding activity could be detected in nuclear extracts of both Jurkat and peripheral blood T cells after simultaneous CD69 and protein kinase C stimulation. Finally, CD69-mediated signals could increase NFAT and AP-1 binding activity following PMA and ionomycin stimulation in peripheral blood T cells. Collectively, these data suggest that CD69-generated signals participate in the control of the IL-2 gene expression at the transcriptional level, likely acting through NFAT and AP-1 transcription factor complexes.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.