Immunological Identification of 1,25-Dihydroxyvitamin D3 Receptors in Human Promyelocytic Leukemic Cells (HL-60) during Homologous Regulation

Immunological techniques were utilized to detect 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) receptor levels and to characterize physical/chemical changes in receptors in human promyelocytic leukemic cells (HL-60) during continuous exposure to hormone. The monoclonal antibody (IVG8C11) raised against th...

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Bibliographic Details
Published in:The Journal of biological chemistry 1989-08, Vol.264 (23), p.13701-13705
Main Authors: Lee, Y, Inaba, M, DeLuca, H F, Mellon, W S
Format: Article
Language:English
Subjects:
Antibodies, Monoclonal
Biological and medical sciences
Calcitriol - metabolism
Calcitriol - pharmacology
Cell Line
Cell Nucleus - drug effects
Cell Nucleus - metabolism
Cell receptors
Cell structures and functions
Cycloheximide - pharmacology
Fundamental and applied biological sciences. Psychology
Humans
Immunoblotting
Kinetics
Leukemia, Promyelocytic, Acute
Miscellaneous
Molecular and cellular biology
Receptors, Calcitriol
Receptors, Steroid - analysis
Receptors, Steroid - drug effects
Receptors, Steroid - metabolism
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Summary:Immunological techniques were utilized to detect 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) receptor levels and to characterize physical/chemical changes in receptors in human promyelocytic leukemic cells (HL-60) during continuous exposure to hormone. The monoclonal antibody (IVG8C11) raised against the porcine intestinal 1,25-(OH)2D3 receptor immunoprecipitated quantitatively 1,25-(OH)2D3 receptors in nuclear extracts from HL-60 cells. The highly enriched immunoprecipitated receptors were electrophoresed on sodium dodecyl sulfate-polyacrylamide gels and transferred to polyvinylidene difluoride membranes, which were probed with 125I-labeled IVG8C11. The basal receptor from the cells treated with 1,25-(OH)2D3 for 2 h was detected as a single form at 53 kDa. Moreover, receptors were shown to be up-regulated at 12 h and down-regulated at 48 and 72 h in the continuous presence of hormone as evidenced by the ratio of density of the bands, 1.0 (2 h):4.2 (12 h):1.2 (48 h):0.9 (72 h), as measured by laser scanning densitometry. The up- and down-regulated receptors were also detected as single forms and had the same molecular mass as the basal receptor. Therefore, the data presented here strongly support the hypothesis of homologous regulation of 1,25-(OH)2D3 receptors in intact human target cells.
ISSN:0021-9258
1083-351X
DOI:10.1016/S0021-9258(18)80055-2