Studies on the structure of the mouse CBF-A gene and properties of a truncated CBF-A isoform generated from an alternatively spliced RNA

• Published in: Gene Vol. 139; no. 2; pp. 147 - 153
• Main Authors: Sohn, Kun-Young, Maity, Sankar N., de Crombrugghe, Benoit
• Format: Journal Article
• Language: English
• Published: Lausanne Elsevier B.V 25.02.1994; Amsterdam Elsevier; New York, NY
• Subjects: Alternative Splicing; Amino Acid Sequence; Animals; Base Sequence; Biological and medical sciences; CCAAT-binding protein; DNA - metabolism; Exons - genetics; Fundamental and applied biological sciences. Psychology; fusion protein; Genes. Genome; Introns - genetics; Mice; Molecular and cellular biology; Molecular genetics; Molecular Sequence Data; Protein Biosynthesis; Recombinant DNA; RNA - genetics; transcription factor; Transcription Factors - genetics; Transcription Factors - metabolism; Transcription, Genetic; aa; transcription factor; ORF; SDS; DTT; CBF; nt; dNTP; GST; AMV; PAGE; Recombinant DNA; bp; ds; tsp; fusion protein; UTR; CCAAT-binding protein; kb; cDNA; oligo; wt; PCR; polymerase chain reaction; Vertebrata; Molecular form; Mammalia; Mouse; Rodentia; Truncated shape; Gene organization; Transcription factor; Structure function relationship
• ISSN: 0378-1119; 1879-0038
• DOI: 10.1016/0378-1119(94)90748-X

CCAAT-binding factor (CBF), a heteromeric transcription factor that binds to sequences containing a CCAAT motif, is composed of three subunits. A, B and C, which are all required for DNA binding. The mouse CBF-A gene contains seven coding exons, which span 12 kb. Evidence is also presented for an additional 5′ untranslated exon. The 90-aminoacid (aa) segment of CBF-A, which shows a high degree of sequence identity with the yeast transcription factor, HAP3, is split into exons 3 and 4. An alternatively spliced RNA that lacks exon 3 was identified by polymerase chain reaction. Although removal of exon 3 interrupts the CBF-A reading frame, a potential start codon at the 3′ end of exon 2 is in the same reading frame as the reading frame encoding CBF-A in exons 4 to 7. A CBF-A polypeptide of the predicted 17-kDa, size, was indeed identified after in vitro transcription and translation of the DNA complementary to RNA (cDNA) corresponding to the alternatively spliced CBF-A mRNA. In contrast to full-length CBF-A, this truncated CBFA did not bind to a DNA sequence containing the CCAAT motif in the presence of the other two components of CBF. This result indicates that the segment corresponding to the exons missing in the truncated isoform of CBF-A is essential for the binding of CBF to DNA.