C/EBP beta (NF-M) is essential for activation of the p20K lipocalin gene in growth-arrested chicken embryo fibroblasts

The p20K gene is induced in conditions of reversible growth arrest in chick en embryo fibroblasts (CEF). This expression is dependent on transcriptiona l activation and on a region of the promoter designated the quiescence-resp onsive unit (QRU). In this report, we describe the regulatory elements of t he QRU responsible for activation in resting cells and characterize the tra ns-acting proteins interacting with these elements. We show that the QRU co nsists of functionally distinct domains including quiescence-specific and w eak proliferation-responsive elements. The quiescence responsiveness of the QRU was mapped to two C/EBP binding sites, and the activity of the p20K pr omoter and its QRU was inhibited by the expression of a dominant negative m utant of C/EBP beta in nondividing cells. The activation of QRU in response to serum starvation and contact inhibition correlated with the presence of a growth arrest-specific complex in electrophoretic mobility shift assays. This complex was supershifted by antibody for C/EBP beta. C/EBP beta accum ulated in conditions of contact inhibition as a result of transcriptional a ctivation. Therefore, C/EBP beta was itself regulated as a growth arrest-sp ecific gene in CEF. Finally, we show that the expression of p20K is regulat ed by linoleic acid, an essential fatty acid binding to p20K. The addition of linoleic acid to contact-inhibited CEF markedly repressed the synthesis of p20K without inducing mitogenesis. The activity of the QRU was inhibited by linoleic acid or the peroxisome proliferator-activated receptor PPAR ga mma 2 in transient expression assays. Therefore, we have identified C/EBP b eta as a key activator of a growth arrest-specific gene in CEF and implicat ed an essential fatty acid, linoleic acid, in regulation of the QRU and the p20K lipocalin gene.