The EGR1 protein contains a discrete transcriptional regulatory domain whose deletion results in a truncated protein that blocks EGR1-induced transcription.
Egr-1 is a ubiquitous immediate-early gene whose expression is induced by a wide range of different stimuli. A requirement for egr-1 expression has been demonstrated in pathways leading to both proliferation and differentiation, suggesting that egr-1 is a critical intermediary in determining the long-term cellular response to a stimulus. To determine how egr-1 coordinates a cellular response to receptor-mediated stimulation, we have developed a transient cotransfection assay to map functional domains in the EGR1 protein. We localized an activation domain to a serine/threonine/proline-rich region between amino acids 174 and 270. Using this information, we designed a mutant that lacks this activation domain, but retains the DNA-binding domain. When cotransfected into fibroblasts with an EGR1-dependent reporter, this mutant inhibited the transcriptional activity of both endogenous EGR1, as well as exogenously expressed, wild-type EGR1 protein. These data demonstrate that the activation domain of EGR1 is critical for the activity of the protein, and that a mutant lacking this domain can dominantly inhibit wild-type EGR1 function.
Carman
J A
JA
Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia 19104, USA.
Monroe
J G
JG
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000032:analysis / Q000096:biosynthesis; D002522:Chlorocebus aethiops; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D051766:Early Growth Response Protein 1; D015870:Gene Expression; D006801:Humans; D017874:Immediate-Early Proteins; D007668:Kidney; D016393:Lymphoma, B-Cell; D051379:Mice; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D017384:Sequence Deletion; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; C486109:Egr1 protein, mouse; D017874:Immediate-Early Proteins; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D014157:Transcription Factors; D015500:Chloramphenicol O-Acetyltransferase