A direct role for c-fos in AP-1-dependent gene transcription.
Transcription factor activator protein 1 (AP-1) is a protein fraction that contains c-fos, c-jun, and several other related proteins. Although this protein fraction can stimulate transcription in vitro, the relative contributions of c-fos and c-jun to the transcriptional effect of AP-1 are not clear. In order to approach this question, we have overexpressed both proteins using a baculovirus-mediated expression system and defined their DNA-binding and transcriptional enhancement activities in vitro. Gel mobility-shift and DNase 1 footprinting assays showed that c-jun protein specifically binds to DNA through an AP-1 binding site. Under the same conditions, no detectable binding of c-fos protein was observed. However, when the DNA binding assays were performed in the presence of both c-jun and c-fos, a marked increase in the affinity of c-jun for the AP-1 site was observed. An AP-1-dependent transcription assay was used to test the capability of both proteins to stimulate correctly initiated RNA synthesis in vitro. Under our conditions, c-jun protein was capable of stimulating specific RNA transcription in an AP-1 site-dependent manner. In contrast, c-fos protein showed no detectable transcriptional activation by itself. However, a transcription assay carried out in the presence of both c-fos and c-jun proteins showed that the c-fos/c-jun complex was more active as a transcriptional regulator than c-jun protein alone. These experimental results indicate that c-fos and c-jun proteins are required to reconstitute full AP-1-dependent transcriptional activation and directly demonstrate that c-fos is a regulator of gene expression.
Herrera
R
R
Dana-Farber Cancer Institute, Boston, Massachusetts 02115.
Agarwal
S
S
Walton
K
K
Satterberg
B
B
Distel
R J
RJ
Goodman
R
R
Spiegelman
B M
BM
Roberts
T M
TM
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000378:metabolism / Q000502:physiology*; D003850:Deoxyribonuclease I; D066298:In Vitro Techniques; D007313:Insecta / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism / Q000502:physiology*; D016760:Proto-Oncogene Proteins c-fos; D016755:Proto-Oncogene Proteins c-jun; D011994:Recombinant Proteins / Q000378:metabolism; D014157:Transcription Factors / Q000502:physiology*; D014158:Transcription, Genetic / Q000235:genetics*
D004268:DNA-Binding Proteins; D011518:Proto-Oncogene Proteins; D016760:Proto-Oncogene Proteins c-fos; D016755:Proto-Oncogene Proteins c-jun; D011994:Recombinant Proteins; D014157:Transcription Factors; D003850:Deoxyribonuclease I