Oxygen stress increases prolyl cis/trans isomerase activity and expression of cyclophilin 18 in rabbit blastocysts.
The peptidyl-prolyl cis/trans isomerase (PPIase) activity and the expression of cyclophilins were studied in 6-day-old rabbit preimplantation embryos cultured under physiological and increased oxygen concentrations of 5% and 20% O(2), respectively. The PPIase activity was completely inhibited by cyclosporin A (CsA). The inhibitor of FK506-binding proteins, rapamycin, had no effect on the PPIase activity, indicating that the PPIase activity in rabbit blastocysts originates from cyclophilins. Using CsA affinity chromatography, only one cyclophilin with a molecular mass of about 17.8 kDa was separated. The cDNA of rabbit cyclophilin was cloned and sequenced. Analysis of the 682-base pair cDNA revealed an open reading frame coding for a polypeptide of 164 amino acid residues with a molecular weight of 17.83 kDa. Homologies of 90% and 96% for the cDNA and amino acid sequence, respectively, to the human CyP18 were found, suggesting that the novel rabbit cyclophilin is a member of the CyP18 family (rabCyP18). The transcription level of rabCyP18 mRNA was 8.3 +/- 0.6 pg in 100 ng total RNA in noncultured blastocysts. In vitro culture with moderate oxygen stress (20% O(2)) resulted in a 1.5-fold increase in rabCyP18 transcription and an increased PPIase activity compared to that of blastocysts cultured with 5% O(2). Increase in transcription rate and PPIase activity by oxygen stress suggests an involvement of CyP18 in oxygen defense in rabbit preimplantation embryos.
Santos
A N
AN
Department of Anatomy and Cell Biology, Martin Luther University Faculty of Medicine, D-06097 Halle (Saale), Germany.
Körber
S
S
Küllertz
G
G
Fischer
G
G
Fischer
B
B
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001755:Blastocyst / Q000378:metabolism*; D003001:Cloning, Molecular; D046508:Culture Techniques; D016572:Cyclosporine / Q000494:pharmacology; D018076:DNA, Complementary / Q000737:chemistry; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015870:Gene Expression; D006801:Humans; D008969:Molecular Sequence Data; D018384:Oxidative Stress; D010100:Oxygen / Q000008:administration & dosage; D019696:Peptidylprolyl Isomerase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D011817:Rabbits; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment
D018076:DNA, Complementary; D004791:Enzyme Inhibitors; D012333:RNA, Messenger; D016572:Cyclosporine; D019696:Peptidylprolyl Isomerase; D010100:Oxygen