Alternative splicing of human prostaglandin G/H synthase mRNA and evidence of differential regulation of the resulting transcripts by transforming growth factor beta 1, interleukin 1 beta, and tumor necrosis factor alpha.
Prostaglandin G/H synthase (PGG/HS) is the rate-limiting enzyme in the conversion of arachidonic acid to prostaglandins and thromboxanes. We screened a human lung fibroblast cDNA library with an ovine PGG/HS cDNA and isolated a 2.3-kilobase clone (HCO-T9). Sequence analysis of this clone showed that (a) it contained the entire translated region of PGG/HS and (b) it displayed an in-frame splicing of the last 111 base pairs encoded by exon 9, which resulted in the elimination of the N-glycosylation site at residue 409. Polymerase chain reaction amplification with specific oligonucleotides of reverse-transcribed mRNA from diverse human tissues and cultured cells yielded 400- and 300-base pair fragments that corresponded, respectively, to the intact and spliced transcripts. The expression of these two transcripts in cultured human lung fibroblasts was differentially regulated by serum, transforming growth factor beta 1, interleukin 1 beta, tumor necrosis factor alpha, and phorbol 12-myristate 13-acetate, as each of these conditions stimulated preferentially the expression of the unspliced transcripts. The elimination of one of the four N-glycosylation sites by the alternative splicing of exon 9 and the differential regulation of this process by relevant cytokines and growth factors may represent a mechanism for the regulation of PGG/HS enzymatic activity under physiological or pathological conditions.
Diaz
A
A
Department of Medicine, Jefferson Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania 19107.
Reginato
A M
AM
Jimenez
S A
SA
D000595:Amino Acid Sequence; D001483:Base Sequence; D015152:Blotting, Northern; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology*; D008168:Lung / Q000166:cytology / Q000378:metabolism; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics* / Q000378:metabolism; D012326:RNA Splicing; D012333:RNA, Messenger / Q000378:metabolism*; D015183:Restriction Mapping; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014158:Transcription, Genetic; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D007375:Interleukin-1; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D014409:Tumor Necrosis Factor-alpha; D004247:DNA; D011451:Prostaglandin-Endoperoxide Synthases; D013755:Tetradecanoylphorbol Acetate