CD2 signalling induces phosphorylation of CREB in primary lymphocytes

Promoter sequences responsive to cyclic AMP (cAMP) are found in a number of cellular genes, and bind transcription factors of the cAMP response elemen t binding protein (CREB)/activating transcription factor-1 (ATF-1) family. We have used a human T-lymphotropic virus type 1 (HTLV-1) model of cAMP res ponse element (CRE) transcription to investigate the influence of lymphocyt e activation on transcription from homologous regions in the viral. promote r. We previously demonstrated increased HTLV-1 transcription following CD2 but not CD3 receptor cross-linking. We hypothesized that this increased vir al transcription was mediated, in part, through the phosphorylation of CREB . Therefore, we investigated CD2 and CD3 receptor-mediated signalling in pr imary human peripheral blood mononuclear cells (PBMC). CD2, but not CD3, cr oss-linking increased cAMP detected by competitive enzyme-linked immunosorb ent assay (ELISA) approximately fourfold. CD2 cross-linking concurrently in creased phosphorylation of CREB detected by immunoblot assay eightfold. Con sistent with post-translational regulation, no change in total level of CRE B protein was observed. Phosphorylation of CREB occurred through a herbimyc in A and Rp-cAMP-sensitive pathway, suggesting phosphorylation required ant ecedent activation of both protein tyrosine kinases (PTK) and protein kinas e A (PKA). Both CD2 and CD3 cross-linking increased binding of nuclear prot eins to a radiolabelled CRE oligonucleotide probe in electrophoretic mobili ty shift assays suggesting that lymphocyte activation enhances binding inde pendently of phosphorylation of CREB at serine 133. These data indicate spe cific modulation of the CREB/ATF-1 family of transcription factors by the C D2 signalling pathway and suggest CD2 receptor modulation of CRE-mediated t ranscription following ligand engagement (e.g. cell-to-cell contact).