Characterization of the rat A2A adenosine receptor gene: a 4.8-kb promoter-proximal DNA fragment confers selective expression in the central nervous system.
We isolated and characterized a 4.8-kb 5' flanking region of the rat A2A adenosine receptor (A2A-R) gene in the present study. Promoter activity was observed with this DNA fragment in PC12 cells and C6 cells which contain endogenous A2A-Rs. A fusion fragment consisting of the 4.8-kb promoter-proximal DNA fragment of the A2A-R gene, and the coding region of lacZ was utilized to produce mice harbouring the fusion gene. In three independent founder lines, proteins and transcripts of the transgene were found in many areas of the central nervous system (CNS), but not in three peripheral tissues examined. Double immunohistochemical analyses revealed that the transgene was coexpressed with endogenous A2A-R and proper neuronal markers in the brain. Specifically, the transgene in the striatum was found in the enkephalin-containing GABAergic neurons and in the cholinergic neurons as was found for the endogenous A2A-R. However, a selectively enriched striatal expression of the transgene was not found as was observed for the endogenous A2A-R. Collectively, the 4.8-kb promoter-proximal DNA fragment of the rat A2A-R gene contains important element(s) to direct its expression in the CNS where functional A2A-R are found, but were not sufficient to confer the highly concentrated expression of the striatal A2A-R. Furthermore, expressions of A2A-R and the transgene were found in both neurons and astrocytes, suggesting that adenosine might mediate its function through A2A-R in both cell types.
Lee
Yi-Chao
YC
Institute of Biomedical Sciences, Academia Sinica, Taipei 11529, Taiwan.
Chien
Chen-Li
CL
Sun
Chung-Nan
CN
Huang
Chuen-Lin
CL
Huang
Nai-Kuei
NK
Chiang
Ming-Chang
MC
Lai
Hsing-Lin
HL
Lin
Yow-Sien
YS
Chou
Szu-Yi
SY
Wang
Chi-Kuang Leo
CK
Tai
Ming-Hong
MH
Liao
Wen-Lin
WL
Lin
Teng-Nan
TN
Liu
Fu-Chin
FC
Chern
Yijuang
Y
D024506:5' Flanking Region; D000262:Adenylyl Cyclases / Q000494:pharmacology; D000818:Animals; D001253:Astrocytes / Q000187:drug effects / Q000378:metabolism; D015139:Blotting, Southern; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002490:Central Nervous System / Q000033:anatomy & histology / Q000187:drug effects / Q000378:metabolism*; D053938:DNA Fragmentation; D015870:Gene Expression / Q000187:drug effects; D017930:Genes, Reporter / Q000502:physiology; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D051379:Mice; D008822:Mice, Transgenic; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D016716:PC12 Cells; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D012313:RNA / Q000378:metabolism; D051381:Rats; D043705:Receptor, Adenosine A2A / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D014162:Transfection; D014404:Tubulin / Q000378:metabolism; D001616:beta-Galactosidase / Q000378:metabolism
D005904:Glial Fibrillary Acidic Protein; D008869:Microtubule-Associated Proteins; D043705:Receptor, Adenosine A2A; D014404:Tubulin; C462941:beta3 tubulin, mouse; D012313:RNA; D001616:beta-Galactosidase; D000262:Adenylyl Cyclases