Two distinct class A helix-loop-helix transcription factors, E2A and BETA1, form separate DNA binding complexes on the insulin gene E box.
Mutations in the RIPE3a element have shown it to be crucial for efficient tissue-specific expression of the insulin gene. In order to isolate factors binding to this element, we used a labeled RIPE3 probe to screen an expression library derived from a hamster insulinoma cell line. We isolated a clone encoding beta-cell E-box transcriptional activator1 (BETA 1). This clone is a member of the class A subfamily of the helix-loop-helix superfamily of transcriptional activators, as determined both by sequence analysis and by functional association with a class B member (myogenin). This clone is related to, but distinct from, other clones isolated from the same library which are also capable of binding RIPE3a. Analysis showed these additional clones to be the hamster homologs of E12 and E47 (German, M. S., Blaner, M. A., Nelson, C., Moss, L. G., and Rutter, W. J. (1991) Mol. Endocrinol. 5, 292-299). Antibodies were raised against BETA 1 and against a common epitope of E12 and E47 to determine which proteins were contained in the native RIPE3a binding complex. Using these antibodies, we were able to separate the complex into major and minor fractions which contained either E12/47 or BETA 1, respectively. Thus, these two gene products are found in separate fractions of the tissue-specific binding activity and are therefore both likely to be important in insulin gene regulation.
Peyton
M
M
Department of Cell Biology, Baylor College of Medicine, Houston, Texas 77030.
Moss
L G
LG
Tsai
M J
MJ
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D051792:Basic Helix-Loop-Helix Proteins; D001667:Binding, Competitive; D002467:Cell Nucleus / Q000378:metabolism; D003001:Cloning, Molecular; D006224:Cricetinae; D018076:DNA, Complementary / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D015723:Gene Library; D005982:Glutathione Transferase / Q000096:biosynthesis / Q000235:genetics; D018257:Helix-Loop-Helix Motifs; D007328:Insulin / Q000235:genetics*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011485:Protein Binding; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013347:Subcellular Fractions / Q000378:metabolism; D051820:TCF Transcription Factors; D015534:Trans-Activators / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D058125:Transcription Factor 7-Like 1 Protein; D014157:Transcription Factors / Q000378:metabolism*
D051792:Basic Helix-Loop-Helix Proteins; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D005982:Glutathione Transferase; D007328:Insulin; D011993:Recombinant Fusion Proteins; D051820:TCF Transcription Factors; D015534:Trans-Activators; D058125:Transcription Factor 7-Like 1 Protein; D014157:Transcription Factors; C089961:BETA1 protein, Cricetinae