Net-b, a Ras-insensitive factor that forms ternary complexes with serum response factor on the serum response element of the fos promoter.
The Ras signalling pathway targets transcription factors such as the ternary complex factors that are recruited by the serum response factor to form complexes on the serum response element (SRE) of the fos promoter. We have identified a new ternary complex factor, Net-b. We report the features of the net gene and show that it produces several splice variants, net-b and net-c. net-b RNA and protein are expressed in a variety of tissues and cell lines. net-c RNA is expressed at low levels, and the protein was not detected, raising the possibility that it is a cryptic splice variant. We have studied the composition of ternary complexes that form on the SRE of the fos promoter with extracts from fibroblasts (NIH 3T3) cultured under various conditions and pre-B cells (70Z/3) before and after differentiation with lipopolysaccharide (LPS). The fibroblast complexes contain mainly Net-b followed by Sap1 and Elk1. Net-b complexes, as well as Sap1 and Elk1, are induced by epidermal growth factor (EGF) stimulation of cells cultured in low serum. Pre-B-cell complexes contain mainly Sap1, with less of Net-b and little of Elk1. There is little change upon LPS-induced differentiation compared to the increase with EGF in fibroblasts. We have also found that Net-b is a nuclear protein that constitutively represses transcription. Net-b is not activated by Ras signalling, in contrast to Net, Sap1a, and Elk1. We have previously reported that down-regulation of Net proteins with antisense RNA increases SRE activity. The increase in SRE activity is observed at low serum levels and is even greater after serum stimulation, showing that the SRE is under negative regulation by Net proteins and the level of repression increases during induction. Net-b, the predominant factor in ternary complexes in fibroblasts, may both keep the activity of the SRE low in the absence of strong inducing conditions and rapidly shut the activity off after stimulation.
Giovane
A
A
Institut de Génétique et de Biologie Moléculaire et Cellulaire, CNRS, INSERM, ULP, Illkirch, France.
Sobieszczuk
P
P
Ayadi
A
A
Maira
S M
SM
Wasylyk
B
B
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes; D002454:Cell Differentiation; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005786:Gene Expression Regulation; D016762:Genes, fos / Q000235:genetics*; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism*; D015513:Oncogene Proteins; D009928:Organ Specificity; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011518:Proto-Oncogene Proteins / Q000032:analysis; D050783:Proto-Oncogene Proteins c-ets; D012326:RNA Splicing; D012333:RNA, Messenger / Q000032:analysis; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D029702:Schizosaccharomyces pombe Proteins; D026362:Serum Response Factor; D015398:Signal Transduction; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D050807:ets-Domain Protein Elk-1; D018631:ras Proteins
D004268:DNA-Binding Proteins; C494113:Elk1 protein, mouse; C086924:Elk3 protein, mouse; D008070:Lipopolysaccharides; D009687:Nuclear Proteins; D015513:Oncogene Proteins; D011518:Proto-Oncogene Proteins; D050783:Proto-Oncogene Proteins c-ets; D012333:RNA, Messenger; D012097:Repressor Proteins; C085981:Sap1 protein, S pombe; D029702:Schizosaccharomyces pombe Proteins; D026362:Serum Response Factor; D014157:Transcription Factors; D050807:ets-Domain Protein Elk-1; D004815:Epidermal Growth Factor; D018631:ras Proteins