TABLE 1.
Relative Binding Affinity of C/EBP Isoforms at Candidate Consensus Sites in the Murine CPO Upstream Gene
| Site Name | Location of Site (nt) (a) | Relative Affinity of Binding to Individual Isoforms (%)(b) | Characteristics of the Binding Site | ||||||
|---|---|---|---|---|---|---|---|---|---|
| C/EBPα | C/EBPβ (LAP) | C/EBPβ (LIP) | C/EBPβ (LAP*) | C/EBPδ | Mean Relative Affinity (%) (c) | Isoform Score (d) | Strength Category (e) | ||
| C1 | −339 | 1.1 | 3.8 | 3.4 | 0.0 | 1.9 | 2.0% | 4 | Moderate |
| C2 | −362 | 0.0 | 0.3 | 0.3 | 0.0 | 0.0 | 0.1% | 2 | Weak |
| C3 | −483 | 9.5 | 7.7 | 10.4 | 0.0 | 4.0 | 6.3% | 4 | Moderate |
| C4 | −570 | 0.5 | 0.8 | 2.6 | 0.0 | 0.0 | 0.8% | 3 | Weak |
| C5 | −631 | 0.0 | 0.0 | 0.0 | 0.0 | 0.0 | 0.0% | 0 | No binding |
| C6 | −647 | 34.4 | 32.2 | 26.6 | 70.8 | 35.5 | 39.9% | 5 | Strong |
| C7 | −754 | 25.4 | 18.4 | 15.0 | 11.4 | 18.9 | 17.8% | 5 | Strong |
| C8 | −836 | 23.4 | 15.4 | 14.8 | 10.4 | 18.2 | 16.5% | 5 | Strong |
| C9 | −910 | 0.0 | 0.0 | 0.0 | 0.0 | 0.0 | 0.0% | 0 | No binding |
| C10 | −930 | 0.9 | 3.6 | 5.7 | 3.1 | 3.8 | 3.4% | 5 | Moderate |
| C11 | −1018 | 0.0 | 3.2 | 2.2 | 3.3 | 5.8 | 2.9% | 4 | Moderate |
| C12 | −1027 | 1.1 | 3.9 | 3.0 | 0.9 | 5.9 | 3.0% | 5 | Moderate |
| C13 | −1096 | 0.0 | 0.4 | 0.0 | 0.0 | 1.0 | 0.3% | 2 | Weak |
| C14 | −1152 | 0.0 | 1.1 | 2.2 | 0.0 | 0.0 | 0.7% | 2 | Weak |
| C15 | −1177 | 0.0 | 1.3 | 2.2 | 0.0 | 0.0 | 0.7% | 2 | Weak |
| C16 | −1220 | 3.5 | 6.7 | 8.0 | 0.0 | 3.8 | 4.4% | 4 | Moderate |
| C17 | −1297 | 0.2 | 1.1 | 3.7 | 0.0 | 1.2 | 1.2% | 4 | Weak |
| Total signal in binding assay: | 100.0 | 100.0 | 100.0 | 100.0 | 100.0 | ||||
Location upstream of ATG in the mCPO promoter. The nucleotide shown is +1 in each 10 bp C/EBP-binding motif as defined in Figure 4.
For each C/EBP protein isoform, binding affinities were estimated from the signal intensity of specific bands on EMSA autoradiograms. (Nuclear proteins from C/EBP-transfected COS-7 cells were bound to individual oligonucleotide probes encompassing each CPO promoter site). For each band, relative affinity was expressed as a percentage of the total signal intensity for that isoform, summed from all 17 promoter sites.
Mean binding affinity, averaged across all five C/EBP isoforms.
Isoform binding score: Number of C/EBP isoforms that showed detectable binding to that particular DNA sequence.
Binding Strength Category of the nucleotide sequence, defined as follows:
| Strong: | Mean binding affinity is > 10%; the site can bind to all five C/EBP isoforms. |
| Moderate: | Mean binding affinity is at least 2%; the site can bind to 4 or more isoforms. |
| Weak: | Mean binding affinity is less than 2%; the site can bind to at least 2 isoforms. |
| Non-binding | The site fails to bind to any of the C/EBP isoforms. |