Design of a C/EBP-specific, dominant-negative bZIP protein with both inhibitory and gain-of-function properties.
We have developed a bZIP protein, GBF-F, with both dominant-negative (DN) and gain-of-function properties. GBF-F is a chimera consisting of two components: the DNA binding (basic) region from the plant bZIP protein GBF-1 (GBF) and a leucine zipper (F) designed to preferentially heterodimerize with the C/EBP alpha leucine zipper. Biochemical studies show that GBF-F preferentially forms heterodimers with C/EBP alpha and thus binds a chimeric DNA sequence composed of the half-sites recognized by the C/EBP and GBF basic regions. Transient transfections in HepG2 hepatoma cells show that both components of GBF-F are necessary for inhibition of C/EBP alpha transactivation. When the C/EBP alpha leucine zipper is replaced with that of either GCN4 or VBP, the resulting protein can transactivate a C/EBP cis-element but is not inhibited by GBF-F, indicating that the specificity of dominant-negative action is determined by the leucine zipper. All known members of the C/EBP family contain similar leucine zipper regions and are inhibited by GBF-F. GBF-F also exhibits gain-of-function properties, since, with the essential cooperation of a C/EBP family member, it can transactivate a promoter containing the chimeric C/EBP/GBF site. This protein therefore has potential utility both as a dominant-negative inhibitor of C/EBP function and as an activator protein with novel DNA sequence specificity.
Olive
M
M
Laboratory of Biochemistry, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Williams
S C
SC
Dezan
C
C
Johnson
P F
PF
Vinson
C
C
D001483:Base Sequence; D050976:Basic-Leucine Zipper Transcription Factors; D022762:CCAAT-Enhancer-Binding Proteins; D017931:DNA Primers / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000737:chemistry*; D051790:G-Box Binding Factors; D005799:Genes, Dominant; D016350:Leucine Zippers; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry*; D011485:Protein Binding; D011993:Recombinant Fusion Proteins; D012045:Regulatory Sequences, Nucleic Acid; D013329:Structure-Activity Relationship; D014157:Transcription Factors; D015533:Transcriptional Activation
D050976:Basic-Leucine Zipper Transcription Factors; D022762:CCAAT-Enhancer-Binding Proteins; D017931:DNA Primers; D004268:DNA-Binding Proteins; D051790:G-Box Binding Factors; D046911:Macromolecular Substances; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors