Cloning of cyclooxygenase-1b (putative COX-3) in mouse.
OBJECTIVE
To clone and sequence cyclooxygenase-1b (COX-1b, also known as COX-3) mRNA and to generate an antibody against the mouse COX-1b protein and to demonstrate its existence in vivo in mouse tissues.


METHODS
10 C57BL/6 mice, 4 COX-1 knockout mice and 4 COX-1 wild type mice were used.


METHODS
COX-1b mRNA sequence was determined by RT-PCR amplification using specific primers followed by DNA sequencing. COX-1b protein expression was determined by Western blotting.


RESULTS
The mouse COX-1b mRNA is a splice variant of the COX-1 mRNA generated by the retention of intron-1. COX-1b mRNA encodes a 127 amino acid protein with no similarity with known COX sequences. We generated an anti-mouse COX-1b antibody and demonstrated the existence of COX-1b protein in vivo with the highest expression in kidney, heart, and neuronal tissues. We also detected COX-1b mRNA and protein expression in COX-1 knockout mice.


CONCLUSIONS
In mouse, COX-1b encodes a protein with a completely different amino acid sequence than COX-1 or COX-2; therefore it is improbable that COX-1b in this species plays a role in prostaglandin-mediated fever and pain. In addition, the COX-1(-/-) mouse is not a COX-1b(-/-) mouse, therefore it cannot be used to elucidate the function of the COX-1b protein.
Kis
B
B
Department of Physiology and Pharmacology, Wake Forest University Health Sciences, Medical Center Blvd., Winston-Salem, NC 27157, USA. bkis@wfubmc.edu
Snipes
J A
JA
Gaspar
T
T
Lenzser
G
G
Tulbert
C D
CD
Busija
D W
DW
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D015971:Gene Expression Regulation, Enzymologic; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism; D017422:Sequence Analysis, DNA; D014018:Tissue Distribution
D012333:RNA, Messenger; D011451:Prostaglandin-Endoperoxide Synthases; C466421:cyclooxygenase-3