Mol. Cell. Biol., Oct 1997, 5667-5678, Vol 17, No. 10
A Giovane, P Sobieszczuk, A Ayadi, SM Maira and B Wasylyk
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.
Copyright © 1997, American Society for Microbiology
Net-b, a Ras-insensitive factor that forms ternary complexes with serum response factor on the serum response element of the fos promoter
Institut de Genetique et de Biologie Moleculaire et Cellulaire, CNRS, INSERM, ULP, Illkirch, France.
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