The constitutive activation of the CEF-4/9E3 chemokine gene depends on C/EBPbeta in v-src transformed chicken embryo fibroblasts.
The CEF-4/9E3 chemokine gene is expressed constitutively in chicken embryo fibroblasts (CEF) transformed by the Rous sarcoma virus (RSV). This aberrant induction is controlled at the transcriptional and post-transcriptional levels. Transcriptional activation depends on multiple elements of the CEF-4 promoter composing a Src-responsive-Unit or SRU. The SRU includes a TPA responsive element, a PRDII/kappaB domain and a CAAT box. In this report, we identify C/EBPbeta as a component of the trans-acting factor interacting with the CAAT box of the CEF-4 promoter. In addition, we show that C/EBPbeta binds to a second element located in proximity of the TRE. A mutation of this distal CAAT box impaired the activation of the CEF-4 promoter by pp60(v-src) indicating that this element is also part of the SRU. Using the RCASBP retroviral vector, we expressed a dominant negative mutant of C/EBPbeta (designated Delta184-C/EBPbeta) in RSV-transformed CEF. Delta184-C/EBPbeta decreased the accumulation of the CEF-4 mRNA and activation of the CEF-4 promoter by pp60(v-src). The induction of the Cox-2 gene (CEF-147) was also reduced by Delta184-C/EBPbeta. The effect of the dominant negative mutant was observed within 1 h of the activation of a thermolabile pp60(v-src) suggesting that C/EBPbeta is an early target of v-src transformation. The dominant negative mutant did not inhibit the transformation of CEF by RSV and in fact accentuated the transformed cell phenotype. Therefore, the activation of C/EBPbeta is important for the expression of v-src regulated genes but is not required for the in vitro transformation of CEF by pp60(v-src).
Gagliardi
M
M
Department of Biology, York University, 4700 Keele Street, Toronto, Ontario M3J 1P3, Canada.
Maynard
S
S
Bojovic
B
B
Bédard
P A
PA
D000818:Animals; D030161:Avian Proteins; D001358:Avian Sarcoma Viruses / Q000235:genetics; D001665:Binding Sites; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000235:genetics* / Q000378:metabolism; D002472:Cell Transformation, Viral / Q000235:genetics*; D002642:Chick Embryo; D016207:Cytokines / Q000235:genetics* / Q000378:metabolism; D005347:Fibroblasts / Q000502:physiology / Q000821:virology; D005786:Gene Expression Regulation; D016391:Genes, src / Q000235:genetics*; D016297:Mutagenesis, Site-Directed; D011401:Promoter Regions, Genetic; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000235:genetics; D020218:Response Elements / Q000235:genetics
D030161:Avian Proteins; D022782:CCAAT-Enhancer-Binding Protein-beta; D016207:Cytokines; C071280:EMF-1 protein, Gallus gallus; D016392:Proto-Oncogene Proteins pp60(c-src)