Characterization of the rat A2a adenosine receptor gene

Ying-Yueh Chu, Ke-Hsin Tu, Y C Lee, Zheng-Jie Kuo, Hsin-Lin Lai, Yijuang Chern

Research output: Contribution to journalArticle

35 Citations (Scopus)

Abstract

To understand the molecular basis for the regulation of rat A2a adenosine receptor (A2a-R) expression, we have characterized the rat A2a-R gene and defined its promoter regions. Through a combination of restriction mapping and sequence analysis, we have demonstrated that the rat A2a-R gene is composed of two exons interrupted by a 7.2-kb intron. Primer extension and RNase protection on RNA isolated from PC12 cells suggested that the A2a-R gene encoded two clusters of alternative transcripts. The most upstream transcription start site was designated as +1. The sequence of the proximal 1.5 kb of 5'-flanking region demonstrated no potential TATA box, CCAAT box, or initiator element in the appropriate location. Varying lengths of 5'-flanking regions were inserted into a transient expression vector (pGL2-basic), which contained bacterial luciferase as the reporter gene, to determine its promoter region(s) in PC12 cells, CHOP cells, and C6 cells. Consistent with two clusters of transcription start sites, two independent functional promoter regions (designated P1, -67/-1; and P2, +272/+304) for the rat A2a-R gene were identified. Although both promoters are in use in PC12 cells, only P2 is active in CHOP cells, indicating possible cell line-specific usage of these two promoters.

Original languageEnglish
Pages (from-to)329-37
Number of pages9
JournalDNA and Cell Biology
Volume15
Issue number4
DOIs
Publication statusPublished - Apr 1996
Externally publishedYes

Fingerprint

Purinergic P1 Receptors
PC12 Cells
Genetic Promoter Regions
Genes
5' Flanking Region
Transcription Initiation Site
Bacterial Luciferases
Restriction Mapping
TATA Box
Ribonucleases
Reporter Genes
Introns
Sequence Analysis
Exons
RNA
Cell Line

Keywords

  • Alternative Splicing
  • Animals
  • Base Sequence
  • Cell Line
  • DNA Primers
  • Dogs
  • Gene Expression Regulation
  • Humans
  • Introns
  • Luciferases
  • Molecular Sequence Data
  • PC12 Cells
  • Promoter Regions, Genetic
  • Rats
  • Receptors, Purinergic P1
  • Recombinant Proteins
  • Regulatory Sequences, Nucleic Acid
  • Sequence Homology, Nucleic Acid
  • Transcription, Genetic
  • Transfection
  • Comparative Study
  • Journal Article
  • Research Support, Non-U.S. Gov't

Cite this

Characterization of the rat A2a adenosine receptor gene. / Chu, Ying-Yueh; Tu, Ke-Hsin; Lee, Y C; Kuo, Zheng-Jie; Lai, Hsin-Lin; Chern, Yijuang.

In: DNA and Cell Biology, Vol. 15, No. 4, 04.1996, p. 329-37.

Research output: Contribution to journalArticle

Chu, Y-Y, Tu, K-H, Lee, YC, Kuo, Z-J, Lai, H-L & Chern, Y 1996, 'Characterization of the rat A2a adenosine receptor gene', DNA and Cell Biology, vol. 15, no. 4, pp. 329-37. https://doi.org/10.1089/dna.1996.15.329
Chu, Ying-Yueh ; Tu, Ke-Hsin ; Lee, Y C ; Kuo, Zheng-Jie ; Lai, Hsin-Lin ; Chern, Yijuang. / Characterization of the rat A2a adenosine receptor gene. In: DNA and Cell Biology. 1996 ; Vol. 15, No. 4. pp. 329-37.
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AB - To understand the molecular basis for the regulation of rat A2a adenosine receptor (A2a-R) expression, we have characterized the rat A2a-R gene and defined its promoter regions. Through a combination of restriction mapping and sequence analysis, we have demonstrated that the rat A2a-R gene is composed of two exons interrupted by a 7.2-kb intron. Primer extension and RNase protection on RNA isolated from PC12 cells suggested that the A2a-R gene encoded two clusters of alternative transcripts. The most upstream transcription start site was designated as +1. The sequence of the proximal 1.5 kb of 5'-flanking region demonstrated no potential TATA box, CCAAT box, or initiator element in the appropriate location. Varying lengths of 5'-flanking regions were inserted into a transient expression vector (pGL2-basic), which contained bacterial luciferase as the reporter gene, to determine its promoter region(s) in PC12 cells, CHOP cells, and C6 cells. Consistent with two clusters of transcription start sites, two independent functional promoter regions (designated P1, -67/-1; and P2, +272/+304) for the rat A2a-R gene were identified. Although both promoters are in use in PC12 cells, only P2 is active in CHOP cells, indicating possible cell line-specific usage of these two promoters.

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