Identification of cytidine deaminase as inhibitor of granulocyte-macrophage colony formation.
Mature human blood granulocytes produce regulatory factors that inhibit colony formation by human and murine granulocyte-macrophage colony-forming cells (GM-CFC). The inhibition of GM-CFC by granulocyte extract (GRE) was strongly enhanced by the addition of thymidine (3 to 6 x 10(-5) M for human cells) and by the presence of fetal calf serum (FCS) in the growth medium. Deoxycytidine and deoxyuridine produced effects similar to those of thymidine, but at higher concentrations (2 to 4 x 10(-4) M). It was further observed that GRE prevented the antiproliferative effects of cytosine arabinoside (Ara-C) and azadeoxycytidine, suggesting that GRE contained cytidine deaminase (CDD) activity, since CDD is known to abolish the effects of these nucleoside analogs. Accordingly, the GRE was tested for and shown to contain an enzymatic activity that converted deoxycytidine to deoxyuridine, confirming the presence of CDD activity in GRE. The GM-CFC inhibition factor was found to copurify with CDD activity during three succeeding chromatographic separations, indicating that CDD was indeed the inhibiting factor itself. This conclusion was further substantiated by gel filtration experiments demonstrating a molecular weight (MW) of approximately 50 kd, which corresponds to the MW previously published for CDD activity. Furthermore, addition of tetrahydrouridine (THU), a known specific inhibitor of CDD, abolished the suppressive effect of GRE on GM-CFC, which independently confirmed the identification of CDD as an inhibitor of GM-CFC. The growth-regulating property of CDD could be explained by depletion of deoxycytidine nucleotides necessary for DNA synthesis or by a direct effect of CDD binding to specific receptors on progenitor cells.
Bøyum
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A
Norwegian Defence Research Establishment, Division for Environmental Toxicology, Kjeller.
Løvhaug
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D
Seeberg
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Nordlie
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EM
D000818:Animals; D001374:Azacitidine / Q000494:pharmacology; D001854:Bone Marrow Cells; D002455:Cell Division / Q000187:drug effects; D003561:Cytarabine / Q000494:pharmacology; D003564:Cytidine Deaminase / Q000378:metabolism*; D004247:DNA / Q000096:biosynthesis; D003841:Deoxycytidine / Q000494:pharmacology; D003857:Deoxyuridine / Q000494:pharmacology; D005260:Female; D006098:Granulocytes / Q000166:cytology* / Q000502:physiology; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D008264:Macrophages / Q000166:cytology*; D051379:Mice; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D013936:Thymidine / Q000494:pharmacology
D003561:Cytarabine; D003841:Deoxycytidine; D004247:DNA; D003564:Cytidine Deaminase; D001374:Azacitidine; D013936:Thymidine; D003857:Deoxyuridine