Identification and characterization of transcriptional arrest sites in exon 1 of the human adenosine deaminase gene.
Analysis of human adenosine deaminase (ADA) gene transcription in four different cell lines indicated that a high density of RNA polymerase II complexes is present at the 5' end of the gene and that the extent of transcription elongation beyond the promoter-proximal region governs gene expression. To determine the sequence requirements for a potential transcription arrest site in the promoter-proximal region, genomic clones containing the ADA promoter, exon 1, and various lengths of intron 1 were injected into Xenopus laevis oocyte germinal vesicles. Transcription analysis indicated that nascent ADA transcripts were highly represented at the promoter-proximal region of the injected templates, suggesting that transcription arrest occurred in the oocyte transcription system. Analysis of the transcription products indicated that ADA transcription initiated at the authentic start site and that the most prominent, short ADA transcripts were 105 nucleotides in length. The 3' end of these transcripts mapped within exon 1, 10 nucleotides downstream of the translation initiation codon. Deletion analysis demonstrated that sequences within exon 1 were sufficient to specify the synthesis of the 105-nucleotide transcripts. Taken together, these data suggest that a transcription arrest mechanism operates in the promoter-proximal region of the human ADA gene and that regulation of elongation beyond this point plays a major role in regulating ADA gene expression.
Verna and Marrs McLean Department of Biochemistry, Baylor College of Medicine, Houston, Texas 77030.
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Chen
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D000243:Adenosine Deaminase / Q000235:genetics*; D000818:Animals; D015152:Blotting, Northern; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D005091:Exons; D005796:Genes; D006801:Humans; D009700:Nucleoside Deaminases / Q000235:genetics*; D009865:Oocytes / Q000201:enzymology; D011401:Promoter Regions, Genetic; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D012319:RNA Polymerase II / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000302:isolation & purification; D015183:Restriction Mapping; D014158:Transcription, Genetic; D014981:Xenopus
D012333:RNA, Messenger; D012313:RNA; D012319:RNA Polymerase II; D009700:Nucleoside Deaminases; D000243:Adenosine Deaminase