DT-diaphorase activity in NSCLC and SCLC cell lines: a role for fos/jun regulation

To assess the potential differential lung tumour expression of NAD(P)H:quinone reductase (NQO1), the human (h) NQO1 promoter was characterized in gene transfer studies. A deletion panel of 5′ flanking hNQO1 promoter constructs was made and tested in transient transfection assays in NSCLC and SCLC cell lines. The largest hNQO1 construct (–1539/+115) containing the antioxidant response element (ARE), exhibited robust levels of reporter activity in the NSCLC (H460, H520, and A549) cell lines and expression was over 12 to 77-fold higher than the minimal (–259/+115) promoter construct. In contrast, there was little difference in promoter activity between the largest and minimal promoter construct in the SCLC (H146, H82 and H187) cell lines. Deletion of the sites for NFκB and AP-2 and the XRE did not significantly affect hNQO1 promoter activity in either the NSCLC or SCLC cell lines. Robust promoter activity in NSCLC lines was mediated by a 359 bp segment of the proximal promoter that contained a canonical AP-1 binding site, TGACTCAG, within the ARE. Gel supershift assays with various specific Fos/Jun antibodies identified Fra1, Fra2 and Jun B binding activity in NSCLC cells to a promoter fragment (–477 to –438) spanning the AP-1 site, whereas SCLC do not appear to express functional Fra or Jun B. These results suggest a possible role for AP-1 activity in the differential expression of hNQO1 in NSCLC. © 1999 Cancer Research Campaig

AP-1 Is a Component of the Transcriptional Network Regulated by GSK-3 in Quiescent Cells

The protein kinase GSK-3 is constitutively active in quiescent cells in the absence of growth factor signaling. Previously, we identified a set of genes that required GSK-3 to maintain their repression during quiescence. Computational analysis of the upstream sequences of these genes predicted transcription factor binding sites for CREB, NFκB and AP-1. In our previous work, contributions of CREB and NFκB were examined. In the current study, the AP-1 component of the signaling network in quiescent cells was explored.Using chromatin immunoprecipitation analysis, two AP-1 family members, c-Jun and JunD, bound to predicted upstream regulatory sequences in 8 of the 12 GSK-3-regulated genes. c-Jun was phosphorylated on threonine 239 by GSK-3 in quiescent cells, consistent with previous studies demonstrating inhibition of c-Jun by GSK-3. Inhibition of GSK-3 attenuated this phosphorylation, resulting in the stabilization of c-Jun. The association of c-Jun with its target sequences was increased by growth factor stimulation as well as by direct GSK-3 inhibition. The physiological role for c-Jun was also confirmed by siRNA inhibition of gene induction.These results indicate that inhibition of c-Jun by GSK-3 contributes to the repression of growth factor-inducible genes in quiescent cells. Together, AP-1, CREB and NFκB form an integrated transcriptional network that is largely responsible for maintaining repression of target genes downstream of GSK-3 signaling

Growth inhibition of non-small cell lung cancer cells by AP-1 blockade using a cJun dominant-negative mutant

cJun, a major constituent of AP-1 transcription factor transducing multiple mitogen growth signals, is frequently overexpressed in non-small cell lung cancers (NSCLCs). The purpose of this study is to determine the effects of AP-1 blockade on the growth of NSCLC cells using a cJun dominant-negative mutant, TAM67. Transiently transfected TAM67 inhibited AP-1 transcriptional activity in NSCLC cell lines, NCI-H1299 (H1299), A549 and NCI-H520 (H520). The colony-forming efficiency of H1299 and A549 was reduced by TAM67, while that of H520 was not. To elucidate the effects of TAM67 on the growth of H1299, we established H1299 clone cells that expressed TAM67 under the control of a doxycycline-inducible promoter. In the H1299 clone cells, the induced TAM67 inhibited anchorage-dependent growth by promoting G1 cell-cycle block, but not by apoptosis. The induced TAM67 decreased the expression of a cell-cycle regulatory protein, cyclin A. TAM67 also inhibited anchorage-independent growth of these cells. Furthermore, TAM67 reduced growth of established xenograft tumours from these cells in nude mice. These results suggest that AP-1 plays an essential role in the growth of at least some of NSCLC cells

Emerging roles of ATF2 and the dynamic AP1 network in cancer

Cooperation among transcription factors is central for their ability to execute specific transcriptional programmes. The AP1 complex exemplifies a network of transcription factors that function in unison under normal circumstances and during the course of tumour development and progression. This Perspective summarizes our current understanding of the changes in members of the AP1 complex and the role of ATF2 as part of this complex in tumorigenesis.Fil: Lopez Bergami, Pablo Roberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental (i); Argentina; ArgentinaFil: Lau, Eric . Burnham Institute for Medical Research; Estados UnidosFil: Ronai, Zeev . Burnham Institute for Medical Research; Estados Unido