The interferon-stimulable response elements of two human genes detect overlapping sets of transcription factors.
We have previously reported three types of DNA-protein complexes, formed specifically with the interferon-stimulable response elements (ISRE) in the 5' flanking DNA of the interferon-inducible 6-16 and 9-27 genes, a type-I interferon-inducible early complex involving factor E (ISGF3), M and G complexes induced more slowly in response to type-I and type-II interferons, respectively and C1/C2, a constitutive complex(s). Similar complexes have been reported by others. The operationally defined band-shift complexes M, G and C1/C2 are shown here to be heterogeneous and to differ in their factor content, depending on the ISRE probe. With a 9-27 ISRE probe the M, G and C1/C2 complexes all contain the gamma subunit of ISGF3, which is present constitutively but is induced in response to IFN-alpha (to yield M) or IFN-gamma (to yield G). In contrast, a 6-16 ISRE probe forms band-shift complexes with IFN-alpha-inducible and IFN-gamma-inducible IRF1 and IRF2. With a 6-16 ISRE probe, therefore, M and G each correspond to two complexes which co-migrate in band-shift assays, one corresponding to IRF1, the other to IRF2. With this probe, the constitutive complex C1/C2 corresponds predominantly to IRF2. Consistent with this, IRF1 and IRF2 have lower affinity for the 9-27 ISRE than the 6-16 ISRE, whereas the reverse is true for E (ISGF3) and its gamma subunit. Relatively small differences in affinity appear sufficient to determine whether or not a band-shift complex is detected. In the case of IRF1 and IRF2, the different affinities for the 6-16 and 9-27 probes are dominated by a dinucleotide sequence in the centre of the 14-nucleotide 'core' ISRE. In contrast, preferential binding of E (ISGF3) by the 39-nucleotide 9-27 ISRE-containing sequence, although ISRE dependent, appears to be mediated by sequences 3' of the 'core' ISRE. Accordingly, these complexes can be simultaneously assayed using a hybrid probe consisting of the 5' flanking region and 'core' ISRE sequences from the 6-16 gene and sequences immediately 3' of the 'core' 9-27 ISRE sequence. No evidence was obtained for a modulatory role in factor binding for a pseudo-ISRE sequence close to ISRE in the 9-27 gene. The precise roles of IRF1 and IRF2 in the induction of IFN-beta and the control of interferon-inducible gene expression remain to be established.(ABSTRACT TRUNCATED AT 400 WORDS)
Parrington
J
J
Imperial Cancer Research Fund Laboratories, London, England.
Rogers
N C
NC
Gewert
D R
DR
Pine
R
R
Veals
S A
SA
Levy
D E
DE
Stark
G R
GR
Kerr
I M
IM
D001483:Base Sequence; D002478:Cells, Cultured; D016384:Consensus Sequence; D015342:DNA Probes / Q000235:genetics; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D050856:Interferon Regulatory Factor-1; D050836:Interferon Regulatory Factor-2; D050832:Interferon-Stimulated Gene Factor 3; D050834:Interferon-Stimulated Gene Factor 3, gamma Subunit; D016898:Interferon-alpha / Q000494:pharmacology; D007371:Interferon-gamma / Q000494:pharmacology; D007372:Interferons / Q000494:pharmacology*; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000096:biosynthesis / Q000235:genetics*; D012045:Regulatory Sequences, Nucleic Acid / Q000187:drug effects / Q000235:genetics*; D012097:Repressor Proteins; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic
D015342:DNA Probes; D004268:DNA-Binding Proteins; C494257:IRF1 protein, human; C494229:IRF2 protein, human; C494227:IRF9 protein, human; D050856:Interferon Regulatory Factor-1; D050836:Interferon Regulatory Factor-2; D050832:Interferon-Stimulated Gene Factor 3; D050834:Interferon-Stimulated Gene Factor 3, gamma Subunit; D016898:Interferon-alpha; D010750:Phosphoproteins; D012097:Repressor Proteins; D014157:Transcription Factors; D007371:Interferon-gamma; D007372:Interferons