Association of the glucocorticoid receptor alternatively-spliced transcript 1A with the presence of the high molecular weight membrane glucocorticoidreceptor in mouse lymphoma cells

Using the combination of a cDNA library prepared from membrane glucocortico id (mGR)-enriched S-49 cells and a mouse leukocyte genomic library, we have cloned a 7.3 kb full-length glucocorticoid receptor 1A cDNA. Primer extens ion, 5'RACE, and long distance PCR identified the transcription start site as being located at 1026 bp from the ATC codon. The first 1,013 nucleotides (nts) of the full length sequence constitute 5' UTR sequence(exon 1), the next 2349 bp, the coding region, and the last 3,907 bp, the 3'UTR. The enti re 5'UTR sequence is unique to transcript 1A. The 3'UTR sequence is similar to 88.5 % conserved with the rat 3'UTR. Western blot analysis compared the molecular weight of in vitro translation products from the cloned 1A cDNA with partially purified cellular mGR. Both preparations contained the novel 150 KD and the 94 KD classical CR peptides, suggesting that transcript 1A encodes both receptor forms. Transfection of mGR-less and glucocorticoid ly sis-resistant AtT-20 and HL-60 cells with full-length GR 1A cDNA imparted b oth mGR expression and glucocorticoid lysis-sensitivity to these cells. (C) 1999 Wiley-Liss, Inc.