Rapid decline in folylpolyglutamate synthetase activity and gene expression during maturation of HL-60 cells. Nature of the effect, impact on folate compound polyglutamate pools, and evidence for programmed down-regulation during maturation.
These studies in HL-60 cells examined the regulation of folylpolyglutamate synthetase (FPGS) activity at the level of gene expression during terminal maturation. Following addition of 210 mM Me2SO to cultures of HL-60 cells at a concentration that induces maturation of 85-90% of the cells, FPGS activity, but not folylpolyglutamate hydrolase (FPGH) activity, was reduced 2-7-fold within 1-5 days. The initial decline in FPGS activity preceded any effect of Me2SO on rate of growth and the increase in appearance of nitro blue tetrazolium-positive cells, a marker of cellular maturation, and the decrease after 5 days of exposure to Me2SO was solely accounted for by a 7-fold decrease in value for Vmax. The same time and concentration dependence for Me2SO was shown for the decline in FPGS activity, increase in nitro blue tetrazolium-positive cells, and decline in the level of a 2.1-kilobase FPGS mRNA during exposure to this inducer. This decline in FPGS mRNA was reversible when Me2SO was removed from the culture medium but only until that time when an appreciable number of cells were committed to terminal maturation. Following growth of HL-60 cells with [3H]MTX, used as a model folate compound, a large reduction in its intracellular polyglutamate pools was shown during maturation which quantitatively reflected the decline in FPGS activity as well as folate transport inward (Sirotnak, F.M., Jacobson, D.M., and Yang, C-H. (1986) J. Biol. Chem. 261, 11150-11156). Other data showed that folate status or obviation of the folate requirement during growth of these cells strongly influenced the rapidity of the onset of maturation following exposure to inducer. Overall, these results show that FPGS activity in HL-60 cells is a marker for proliferative capacity and that the underlying basis for the decline in FPGS activity during maturation is altered cognate gene expression which is manifested as early reversible and late irreversible phases. They also suggest that the coordinate reduction observed in folate transport, FPGS activity, dihydrofolate reductase, and probably other folate related enzymes by limiting macromolecular biosynthesis may be early programmed events in the maturation process that influence the switch from proliferation to senescence in these cells.
Egan
M G
MG
Program in Molecular Pharmacology and Therapeutics, Memorial Sloan-Kettering Cancer Center, New York, New York.
Sirlin
S
S
Rumberger
B G
BG
Garrow
T A
TA
Shane
B
B
Sirotnak
F M
FM
D015152:Blotting, Northern; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D004121:Dimethyl Sulfoxide / Q000494:pharmacology*; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D019271:Hypoxanthine; D007042:Hypoxanthines / Q000494:pharmacology; D007700:Kinetics; D015473:Leukemia, Promyelocytic, Acute; D008727:Methotrexate / Q000378:metabolism* / Q000494:pharmacology; D010453:Peptide Synthases / Q000096:biosynthesis / Q000378:metabolism*; D011624:Pteroylpolyglutamic Acids / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis; D013936:Thymidine / Q000494:pharmacology; D013997:Time Factors; D014407:Tumor Cells, Cultured; D011623:gamma-Glutamyl Hydrolase / Q000378:metabolism
D007042:Hypoxanthines; D011624:Pteroylpolyglutamic Acids; D012333:RNA, Messenger; D019271:Hypoxanthine; D011623:gamma-Glutamyl Hydrolase; D010453:Peptide Synthases; C023103:folylpolyglutamate synthetase; D013936:Thymidine; D008727:Methotrexate; D004121:Dimethyl Sulfoxide