The factor binding to the glucocorticoid modulatory element of the tyrosine aminotransferase gene is a novel and ubiquitous heteromeric complex.
Glucocorticoid induction of the tyrosine aminotransferase gene deviates from that of many glucocorticoid-responsive genes by having a lower EC50 and displaying more agonist activity with a given antiglucocorticoid. A cis-acting element, located 3646 base pairs upstream of the start of tyrosine aminotransferase gene transcription, has been found to be sufficient to reproduce these variations with heterologous genes and promoters (Oshima, H., and Simons, S.S., Jr. (1992) Mol. Endocrinol. 6, 416-428). This element has been called a glucocorticoid modulatory element, or GME. Others have called this sequence a cyclic AMP-responsive element (CRE) due to the binding of the cyclic AMP response element binding protein (CREB). We now report the partial purification and characterization of two new proteins (GMEB1 and -2) of 88 and 67 kDa that bind to the GME/CRE as a heteromeric complex. This purification was followed by the formation of a previously characterized, biologically relevant band in gel shift assays. By several biochemical criteria, the GMEBs differed from many of the previously described CREB/CREM/ATF family members. Partial peptide sequencing revealed that the sequences of these two proteins have not yet been described. Size exclusion chromatography and molecular weight measurements of the gel-shifted band demonstrated that the GMEBs bound to the GME as a macromolecular complex of about 550 kDa that could be dissociated by deoxycholate. Similar experiments showed that CREB bound to the GME as heteromeric complexes of about 310 and 360 kDa. As determined from gel shift assays, GMEB1 and -2 are not restricted to rat liver cells but appear to be ubiquitous. Thus, these novel GMEBs may participate in a similar modulation of other glucocorticoid-inducible genes in a variety of cells.
Oshima
H
H
Steroid Hormones Section, NIDDK/Laboratory of Molecular and Cellular Biology, National Institutes of Health, Bethesda, Maryland 20892, USA.
Szapary
D
D
Simons
S S
SS
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002467:Cell Nucleus / Q000378:metabolism; D002846:Chromatography, Affinity; D017362:Cyclic AMP Response Element-Binding Protein / Q000096:biosynthesis / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D005938:Glucocorticoids / Q000378:metabolism* / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D007739:L Cells; D008114:Liver Neoplasms, Experimental; D046911:Macromolecular Substances; D008745:Methylation; D051379:Mice; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D014176:Protein Biosynthesis; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D014157:Transcription Factors / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured; D014444:Tyrosine Transaminase / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification
D017362:Cyclic AMP Response Element-Binding Protein; C490664:GMEB1 protein, human; D005938:Glucocorticoids; C095568:Gmeb1 protein, mouse; C490662:Gmeb2 protein, mouse; D046911:Macromolecular Substances; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D014157:Transcription Factors; D014444:Tyrosine Transaminase