The dbl oncogene is generated by substitution of the 5' portion of its normal counterpart with an unrelated human sequence. To analyze the genomic structure and transcriptional regulation of the dbl proto-oncogene, we have isolated human genomic clones containing the entire human proto-dbl gene, localized in Xq26. Restriction mapping of a 600 kb YAC clone (yWXD311) placed proto-dbl about 50 kb telomeric to the coagulation Factor IX gene. The genomic DNA fragment containing the 5' end of proto-dbl was subcloned into plasmid vectors and the nucleotide sequences of exon I, the flanking intronic region and genomic DNA 5' of the first codon were determined. Sequence analysis of 85119 bp from the region revealed the genomic structure of proto-dbl. It contains 25 exons coding for a 4.7 kb transcript including large 5'- and 3'- (1218 bp and 701 bp, respectively) untranslated regions (UTRs). RNase protection and primer extension assays on RNA from medullary thyroid carcinoma (TT) cells, which normally express dbl, revealed a transcription start site 1218 bp upstream of the ATG of the first exon, A 1.6 kb genomic 5' of the translation start sites drives the expression of a CAT-reporter in transient transfections in the TT cell line, though lacking TATA or CAAT boxes. (C) 2000 Elsevier Science B.V. All rights reserved.
Human dbl proto-oncogene in 85 kb of Xq26, and determination of the transcription initiation site
Ciccodicola A
2000-01-01
Abstract
The dbl oncogene is generated by substitution of the 5' portion of its normal counterpart with an unrelated human sequence. To analyze the genomic structure and transcriptional regulation of the dbl proto-oncogene, we have isolated human genomic clones containing the entire human proto-dbl gene, localized in Xq26. Restriction mapping of a 600 kb YAC clone (yWXD311) placed proto-dbl about 50 kb telomeric to the coagulation Factor IX gene. The genomic DNA fragment containing the 5' end of proto-dbl was subcloned into plasmid vectors and the nucleotide sequences of exon I, the flanking intronic region and genomic DNA 5' of the first codon were determined. Sequence analysis of 85119 bp from the region revealed the genomic structure of proto-dbl. It contains 25 exons coding for a 4.7 kb transcript including large 5'- and 3'- (1218 bp and 701 bp, respectively) untranslated regions (UTRs). RNase protection and primer extension assays on RNA from medullary thyroid carcinoma (TT) cells, which normally express dbl, revealed a transcription start site 1218 bp upstream of the ATG of the first exon, A 1.6 kb genomic 5' of the translation start sites drives the expression of a CAT-reporter in transient transfections in the TT cell line, though lacking TATA or CAAT boxes. (C) 2000 Elsevier Science B.V. All rights reserved.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
