TRANSCRIPTIONAL REGULATION OF INTERCELLULAR-ADHESION MOLECULE-1 BY PHORBOL ESTER IN HUMAN NEUROBLASTOMA CELL-LINE SK-N-SH INVOLVES JUN-CONTAINING AND FOS-CONTAINING ACTIVATOR PROTEIN-1 SITE BINDING COMPLEX(ES)

In this study, the regulatory elements involved in ICAM-1 transcriptio nal response to phorbol ester (12-O-tetradecanoylphorbol-13-acetate; T PA) have been investigated in the human neuroblastoma cell line, SK-N- SH. TPA induced intercellular adhesion molecule 1 (ICAM-1) protein exp ression in SK-N-SH cells within 24 h of treatment as judged by indirec t immunofluorescence. nasal ICAM-1 mRNA levels were barely detectable in untreated SK-N-SH cells but were induced by TPA to a maximal level within 4 h and were reduced thereafter, Analysis of the 5' promoter se quence of ICAM-1 revealed two regions that functioned equally in the T PA induction of ICAM-1 transcription. The first region (-145 to -227) contained a nuclear factor-kappa B (NF kappa B) element, The second re gion (-316 to -390) contained a putative TPA-responsive element (TRE; TGATTCA) and a TATA box. Deletion and point mutation of the latter reg ion indicated that the TRE was indeed the functional element within th is region and acted fully and independently of all other elements incl uding the TATA box at position -352, This TRE bound TPA induced specif ic nuclear complexes in vitro containing junD, c-jun, c-fos, and fra2 but not cAMP-responsive element binding/activating transcription facto r family proteins. ICAM-TRE binding activity was induced within 30 min following TPA treatment. This preceded the appearance of ICAM-NF kapp a B site binding activity. Cotransfection of c-jun and c-fos expressio n vectors into SK-N-SH cells induced transactivation from ICAM-1 promo ter constructs containing the intact but not mutated TRE site. Primer extension analyses revealed that TPA had induced transcription exclusi vely at two sites -40 and -41 bp upstream of the translation start sit e, These data show that the ICAM-TRE and its cognate jun- and fos-cont aining transcription factors play a predominant role in the transcript ional response of ICAM-1 to the protein kinase C activator TPA in SK-N -SH cells.