Transcriptional and post-transcriptional regulation of κB-controlled genes by pp60^v-src

Abstract

The CEF-4/9E3 gene is expressed aberrantly in chicken embryo fibroblasts transformed by the Rous sarcoma virus. This aberrant expression is dependent on transcriptional activation and on the stabilization of the CEF-4 mRNA. The characterization of the CEF-4 promoter indicated that three distinct regulatory elements corresponding to an AP-1 binding site, a PRDII/κB domain and a CAAT box are involved in the activation by pp60^v-src. Several v-src responsive genes are controlled by AP-1 and members of the Ets family but few appear to be dependent on NF-κB. In this study we characterize the expression of genes regulated by NF-κB in normal and RSV-transformed CEF. Run-on transcription analysis indicated that pp60^v-src induces the transcription of several genes controlled by NF-κB but at different levels. While the transcription of CEF-4 was strongly stimulated, that of NF-κB1, c-rel, p53 or IκB-α was activated more modestly by pp60^v-src. In addition the CEF-4 mRNA was the only mRNA species to accumulate significantly in transformed CEF. The ectopic expression of RelA or Rel resulted in the stimulation of the transcription of several known targets of NF-κB. However, the mRNA for IκB-α was the only mRNA species to accumulate considerably in the RelA- or Rel-expressing cells. Hence for most κB-controlled genes, transcriptional activation was not sufficient to obtain a significant increase in mRNA expression. Likewise, RelA or Rel enhanced the transcription of the CEF-4 gene without a significant accumulation of the CEF-4 mRNA. However, transformation by v-src caused a massive accumulation of the CEF-4 mRNA but not of other mRNA species in the RelA- and Rel-expressing cells. Transient expression assays, run-on transcription and Northern blotting analyses indicated that the effect of pp60^v-src on CEF-4 expression was mediated predominantly at the post-transcriptional level in these cells. Therefore transcriptional and post-transcriptional mechanisms determine the restricted pattern of activation of κB-controlled genes in RSV-transformed CEF.