Cloning and characterization of a growth factor-inducible cyclooxygenase gene from rat intestinal epithelial cells.
Growth factors have been shown to play a role in intestinal epithelial growth regulation and transformation. Utilizing standard differential cloning techniques, we have isolated a growth factor-inducible gene (RS-2) from rat intestinal epithelial cells that has approximately 95% homology to the mouse mitogen-inducible cyclooxygenase (COX-2) at the amino acid level. This cDNA hybridizes to a approximately 4.5-kb mRNA from transforming growth factor (TGF)-alpha-stimulated rat intestinal epithelial (RIE-1) cells and is constitutively expressed in vivo in adult rat kidney and brain. Nuclear run-on experiments demonstrate that the increase of RS-2 mRNA after TGF-alpha stimulation is in part due to an increased transcription rate of the gene. The coding region for RS-2 was subcloned into a pCMV-2 expression vector, and the RS-2 protein was expressed in COS-1 cells. Microsomal fractions isolated from the COS-1 cells transfected with the RS-2 expression vector contained cyclooxygenase activity. In addition to the production of prostaglandins, the recombinant RS-2 protein also catalyzed the formation of three other eicosanoid products. In summary, we have cloned a mitogen-inducible cyclooxygenase gene from rat intestinal cells that is induced following growth factor stimulation.
DuBois
R N
RN
Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee.
Tsujii
M
M
Bishop
P
P
Awad
J A
JA
Makita
K
K
Lanahan
A
A
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D001921:Brain / Q000201:enzymology; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000032:analysis; D004790:Enzyme Induction; D004848:Epithelium / Q000187:drug effects / Q000201:enzymology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D015723:Gene Library; D007422:Intestines; D007668:Kidney / Q000201:enzymology; D051379:Mice; D008861:Microsomes / Q000201:enzymology; D008969:Molecular Sequence Data; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis*; D012333:RNA, Messenger / Q000096:biosynthesis* / Q000378:metabolism; D051381:Rats; D017386:Sequence Homology, Amino Acid; D014162:Transfection; D016211:Transforming Growth Factor alpha / Q000494:pharmacology*
D018076:DNA, Complementary; D012333:RNA, Messenger; D016211:Transforming Growth Factor alpha; D011451:Prostaglandin-Endoperoxide Synthases