Cloning of two novel forms of human acidic fibroblast growth factor (aFGF) mRNA
| Autor: | Stephen E. Harris, Halit Canatan, Robert A. Payson, Wen-Pin Wang, Rene L. Myers, Maqsood A. Chotani, Ing-Ming Chiu |
|---|---|
| Rok vydání: | 1993 |
| Předmět: |
Messenger RNA
Reporter gene Base Sequence Molecular Sequence Data Promoter DNA Biology Polymerase Chain Reaction Molecular biology Reverse transcriptase Chloramphenicol acetyltransferase genomic DNA Cricetinae Sequence Homology Nucleic Acid Complementary DNA Gene expression Tumor Cells Cultured Genetics Animals Fibroblast Growth Factor 1 Humans RNA Messenger Cloning Molecular |
| Zdroj: | Nucleic Acids Research. 21:489-495 |
| ISSN: | 1362-4962 0305-1048 |
| DOI: | 10.1093/nar/21.3.489 |
| Popis: | We have previously isolated two different aFGF cDNA clones from kidney and brain. The two corresponding mRNA, designated aFGF 1.A and 1.B, are the predominant species in kidney and brain, respectively. During the characterization of aFGF mRNA in glioblastoma cells, we demonstrated that aFGF mRNA in U1242MG and D65MG glioblastoma cells contain 5'-untranslated sequences different from those of 1.A and 1.B. Through a strategy combining chromosome walking, identification and sequencing of evolutionarily conserved DNA regions, and a reverse transcription and polymerase chain reaction (RT-PCR)-based assay for RNA expression, we have isolated two novel aFGF cDNA clones. The cDNA clone representing aFGF mRNA 1.C was isolated from U1242MG cells; another aFGF cDNA, designated 1.D, was isolated from D65MG cells. Promoter 1C has extensive sequence homology to the hamster aFGF gene promoter that was shown to respond to testosterone stimulation by chloramphenicol acetyltransferase reporter gene assays. Using RT-PCR, we showed that normal, benign and cancerous prostate tissues do not express aFGF 1.C mRNA. In contrast, a prostate carcinoma cell line (PC-3) expresses 1.C mRNA. RT-PCR using 1.D-specific primers showed that kidney, brain and prostate do not express 1.D mRNA even though kidney and brain are the most abundant source for aFGF protein. RNase protection analysis further showed that 1.D mRNA is the predominant aFGF transcript in D65MG glioblastoma cells and in NFF-6 neonatal foreskin fibroblast cells. The genomic DNA corresponding to these two cDNA clones and the 5'-flanking regions were also isolated and their sequences determined. These DNA clones will provide important reagents for studying the regulatory elements of aFGF gene expression. |
| Databáze: | OpenAIRE |
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