Identification of cAMP analogue inducible genes in RAW264 macrophages.
RNA was isolated from RAW264 cells treated with or without 8-Br-cAMP and the differential display and subtractive hybridization methods were performed. One hundred and twenty-five differentially displayed bands were identified. Upon Northern blot analysis, only three of these bands were confirmed as cAMP inducible mRNAs, named cI-1, cI-2, and cI-3 (for cAMP inducible genes 1-3). The cI-3 probe was identical to a previously known gene, gly96. Using the novel cI-1 and cI-2 partial cDNAs as probes, a mouse macrophage cDNA library was screened and the two full length genes were cloned, sequenced, and characterized as encoding large hydrophobic proteins. One hundred and fifteen partial cDNA clones from a subtractive hybridization library were also screened by Northern blot and 64 were found to be cAMP inducible. Of these, 45 represented 31 known unique genes in the GenBank nr database (cI-4-34), and 19 clones representing 15 unique sequences were not in the nr database (cI-35-49). One of the previously known genes was ABC1, the Tangier disease gene, which was identified from four independent partial cDNAs. ABC1 was upregulated in RAW cells by cAMP, concurrent with the cAMP induction of lipid efflux to apolipoprotein A1.
Takahashi
Y
Y
Laboratory of Biochemical Genetics and Metabolism, Box 179, The Rockefeller University, 1230 York Avenue, New York, NY 10021, USA.
Miyata
M
M
Zheng
P
P
Imazato
T
T
Horwitz
A
A
Smith
J D
JD
D000818:Animals; D016632:Apolipoprotein A-I / Q000378:metabolism; D051017:Apoptosis Regulatory Proteins; D001692:Biological Transport; D015152:Blotting, Northern; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000494:pharmacology*; D018076:DNA, Complementary / Q000032:analysis; D005786:Gene Expression Regulation / Q000187:drug effects*; D006824:Hybridization, Genetic; D017874:Immediate-Early Proteins; D050356:Lipid Metabolism; D008264:Macrophages / Q000187:drug effects* / Q000502:physiology; D008565:Membrane Proteins; D051379:Mice; D009363:Neoplasm Proteins; D011487:Protein Conformation; D013997:Time Factors
D016632:Apolipoprotein A-I; D051017:Apoptosis Regulatory Proteins; D018076:DNA, Complementary; C099815:IER3 protein, human; D017874:Immediate-Early Proteins; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D000242:Cyclic AMP